De novo transcriptome assembly: A comprehensive cross-species comparison of
short-read RNA-Seq assemblers

Supplementary Data

Martin Hölzer and Manja Marz

Contents

• S1: Data sets and preprocessing
• S2: Assembly tools
• S3: Executed assembly commands
• S4: HISAT2 re-mapping rate
• S5: RNAQuast statistics
• S6: TransRate
• S7: ExN50
• S8: BUSCO
• S9: DETONATE
• S10: Selected main metrics
• S11: Runtime and memory consumption
• S12: (0,1)-normalized scores per data set and metric

All big data files (processed read data, assemblies, ...) as well as execution commands and Blast results are also available at the Open Science Framework under accession doi.org/10.17605/OSF.IO/5ZDX4.

S1: Data sets and preprocessing

Overview of all data sets used for assembly. All reads were quality checked with FASTQC and Prinseq prior assembly. Shown are read statistics before and after trimming. If a data set was sequenced by preserving the strand specificity (ss), this is indicated by the direction of the sequenced read (pairs): F -- forward, R -- reverse.

ID	Species	Domain	Encoding	Type	Stranded	# Reads (raw)	Read length (raw)	%GC (raw)	FastQC (raw)	# Reads (trimmed)	Read length (trimmed)	%GC (trimmed)	FastQC (trimmed)
ECO	Escherichia coli	Bacteria	Illumina 1.9	single-end	ss F	7943130	94	40%	SINGLE	6643709	25-94	50%	SINGLE
CAL3	Candida albicans	Fungi	Illumina 1.9	paired-end	not	11576932	51	38%	FORWARD | REVERSE	11168529	25-51	38%	FORWARD | REVERSE
ATH	Arabidopsis thaliana	Plant	Illumina 1.9	single-end	not	16911774	30-101	46%	SINGLE	16828036	25-101	46%	SINGLE
MMU	Mus musculus	Mammal	Illumina 1.9	paired-end	ss FR	52645238	76	48%	FORWARD | REVERSE	43321537	25-76	48%	FORWARD | REVERSE
HSA	Homo sapiens	Mammal	Illumina 1.9	paired-end	ss FR	97548052	101	53%	FORWARD | REVERSE	96093116	25-101	52%	FORWARD | REVERSE
EBOV_HSA_3H	Homo sapiens + EBOV 3h	Mammal+Virus	Illumina 1.9	paired-end	not	17203785	20-100	48%	FORWARD | REVERSE	15649763	25-100	47%	FORWARD | REVERSE
EBOV_HSA_7H	Homo sapiens + EBOV 7h	Mammal+Virus	Illumina 1.9	paired-end	not	24688523	20-100	47%	FORWARD | REVERSE	22268601	25-100	47%	FORWARD | REVERSE
EBOV_HSA_23H	Homo sapiens + EBOV 23h	Mammal+Virus	Illumina 1.9	paired-end	not	26470155	20-100	46%	FORWARD | REVERSE	23930250	25-100	46%	FORWARD | REVERSE
HSA_FLUX	Homo sapiens simulated	Mammal-simulated	Illumina 1.9	paired-end	not	60002178	3-100	50%	FORWARD | REVERSE	56962181	25-100	49%	FORWARD | REVERSE

S2: Assembly tools

Overview about the evaluated assembly tools. We obtained the most recent versions in November 2018. We chose Oases and Trans-ABySS as two transcriptome assemblers build on top of the genome assemblers Velvet and ABySS, respectively. To achieve an assembly with Oases or Trans-ABySS one first has to run the underlying genome assembly tool with a range of different k-mers. SOAPdenovo-Trans, build on the principles of SOAPdenovo2, can be used as a stand alone tool for de novo transcriptome assembly based on single k-mer values. Trinity was also especially designed for transcriptome assembly, using only a fixed k-mer value of 25. All these four transcriptome assemblers are designed in the scope of working with RNA-Seq data and are based on de Bruijn graph algorithms. In contrast the Mira assembler works on overlap- graphs and therefore uses no k-mer approach to decompose reads before assembly. It can be run in EST mode to deal with the special requirements of RNA-Seq data. Next to this transcriptome assemblers we also chose one de novo genome assembly tool, SPAdes, originally developed for smaller bacterial-size genomes and single-cell data and also based on de Bruijn graph and multiple k-mer values. MK – Yes, if the tool has an build in multiple kmer approach and automatically merges the output of different kmer runs. ^aOases was used on top of the de novo genome assembler Velvet (v1.2.10). ^bSPAdes, originally designed as a de novo genome assembler for single-cell data, was used in RNA-Seq modus (-rna) and single-cell modus (-sc), respectively. ^cWhen running SPAdes in RNA-Seq modus, two k-mer values are used.

Assembler	Version	Multiple k-mer mode	PMID
Trinity	2.8.4	no	21572440
Trans-ABySS	2.0.1	yes	20935650
SOAPdenovo-Trans	1.03	no	24532719
Oasesa	0.2.08	yes	22368243
IDBA-Tran	1.1.1	yes	23813001
Bridger	v2014-12-01	no	25723335
BinPacker	1.0	no	26894997
Shannon	0.0.2	no	bioRxiv
SPAdes-scb	3.13.0	yes	22506599
SPAdes-rnab	3.13.0	yesc	bioRxiv

S3: Executed assembly commands

In the following we provide all executed commands that were used to run the different assemblies. Each assembler listed in Tab. S2 was run on each data set listed in Tab. S1. For details like used k-mers and strand-specificity download the readme files or expand the view for the corresponding data set. The command files still include the execution command of MIRA, however the output was not used in the comparison because MIRA performed worse on all data sets (very short contigs, almost no homology to known transcripts, bad re-mapping rate, ... see manuscript).

Assembly of Escherichia coli (download script)


#######################################
## Eco1 1x 94bp, strand specific F
#######################################

##PARAMETERS SHARED
PROJECT=eco
R1=eco1_formated.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=35
kmer3=45
kmer4=55
kmer5=65


###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --single $R1 --CPU $THREADS --output $DIR/trinity/ --SS_lib_type F
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --se $R1 --SS --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer5 --SS --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fa
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/$PROJECT.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=94" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=94" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q="$R1 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with default settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/$PROJECT.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades
rnaspades.py --s1 $R1 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc --s1 $R1 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=10
type=-short

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 -strand_specific " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --filter $R1 $DIR/idba-tran/r1.fasta
idba_tran -r $DIR/idba-tran/r1.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer5 --step 10 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fa
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --single $R1 --SS_lib_type F --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --single $R1 --SS_lib_type F --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
cd $OUT
BinPacker -s fq -p single -u $R1 -m F -o $OUT
cd ..
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --single $R1 --ss
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = se" >> $DIR/mira/manifest.config
echo "data = $R1" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
#echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################




Assembly of Candida albicans (download script)


#######################################
## CAL, 2x 51bp not strand-specific
#######################################

##PARAMETERS SHARED
PROJECT=cal3
R1=cal3_1_formated.fastq
R2=cal3_2_formated.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=21
kmer2=27
kmer3=33
kmer4=39

###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer4 --SS --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=51" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=51" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES

##ATTENTION CHANGED KMER BECAUS DEFAULT 55 IS TO BIG
mkdir -p $DIR/rnaspades
rnaspades.py -k 25 --pe1-1 $R1 --pe1-2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc --pe1-1 $R1 --pe1-2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
##################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer4 + 1))
k_step=6
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer4 --step 6 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -o $OUT
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################




Assembly of Arabidopsis thaliana (download script)


#######################################
## Arabidopsis 1x 100bp, not strand specific
#######################################

##PARAMETERS SHARED
PROJECT=ath
R1=ath_formated.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=35
kmer3=45
kmer4=55
kmer5=65

###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --single $R1 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --se $R1 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer5 --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/$PROJECT.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq single end files" >> $CONFIG
echo "q="$R1 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with default settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/$PROJECT.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades
rnaspades.py --s1 $R1 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc --s1 $R1 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=10
type=-short

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --filter $R1 $DIR/idba-tran/r1.fasta
idba_tran -r $DIR/idba-tran/r1.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer5 --step 10 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --single $R1 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --single $R1 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
cd $OUT
BinPacker -s fq -p single -u $R1 -o $OUT
cd ..
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --single $R1
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = se" >> $DIR/mira/manifest.config
echo "data = $R1" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################




Assembly of Mus musculus (download script)


#######################################
## MMU TRINITY TEST DATA, 2x 76bp, strand specific RF
#######################################

##PARAMETERS SHARED
PROJECT=mmu
R1=mmu_1_formated.fastq
R2=mmu_2_formated.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=35
kmer3=45
kmer4=55

###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/ --SS_lib_type FR
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --SS --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer4 --SS --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=76" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=76" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=1" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer4

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades
rnaspades.py --pe1-1 $R1 --pe1-2 $R2 --pe1-fr -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc --pe1-1 $R1 --pe1-2 $R2 --pe1-fr -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
##################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer4 + 1))
k_step=10
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -strand_specific -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
R1=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/mmu_1_formated_reversed.fastq
R2=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/mmu_2_formated_reversed.fastq
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer4 --step 10 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
R1=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/mmu_1_formated.fastq
R2=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/mmu_2_formated.fastq
###################################################################################

###################################################################################
## BRIDGER
#Error! try unstranded mode!
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -m RF -o $OUT
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2 --ss
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
#echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config
echo "segment_placement = <--- --->" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################




Assembly of Homo sapiens (download script)


#######################################
## HSA SRA, 2x 100bp, strand specific RF
#######################################

##PARAMETERS SHARED
PROJECT=hsa
R1=hsa_1.fastq
R2=hsa_2.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=35
kmer3=45
kmer4=55
kmer5=65

###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/ --SS_lib_type FR
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --SS --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer5 --SS --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=1" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades
rnaspades.py --pe1-1 $R1 --pe1-2 $R2 --pe1-fr -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc --pe1-1 $R1 --pe1-2 $R2 --pe1-fr -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=10
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -strand_specific -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
R1=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/hsa_1_reversed.fastq
R2=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/hsa_2_reversed.fastq
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer5 --step 10 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
R1=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/hsa_1.fastq
R2=/mnt/dessertlocal/projects/transcriptome_assembly/data/prinseq/hsa_2.fastq
###################################################################################

###################################################################################
## BRIDGER
# Error, try unstranded
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -m RF -o $OUT
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python /mnt/prostlocal/programs/shannon/0.0.2/shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2 --ss
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
#echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config
echo "segment_placement = <--- --->" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################




Assembly of Homo sapiens + EBOV 3h (download script)


#######################################
## EBOV HSA 3h
## paired-end, not strand specific, 100bp
#######################################

##PARAMETERS SHARED
PROJECT=ebov_hsa_3h
R1=ebov_hsa_3h_1.fastq
R2=ebov_hsa_3h_2.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=29
kmer3=33
kmer4=37
kmer5=41


###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink 25 --maxk 41 --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades/
rnaspades.py -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=4
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink 25 --maxk 41 --step 4
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
cd $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -o $DIR/binpacker
cd ..
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
###################################################################################





Assembly of Homo sapiens + EBOV 7h (download script)


#######################################
## EBOV HSA 7h
## paired-end, not strand specific, 100bp
#######################################

##PARAMETERS SHARED
PROJECT=ebov_hsa_7h
R1=ebov_hsa_7h_1.fastq
R2=ebov_hsa_7h_2.fastq
THREADS=48
MEM=400

DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=29
kmer3=33
kmer4=37
kmer5=41


###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink 25 --maxk 41 --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades/
rnaspades.py -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=4
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink 25 --maxk 41 --step 4
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
cd $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -o $DIR/binpacker
cd ..
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
###################################################################################




Assembly of Homo sapiens + EBOV 23h (download script)


#######################################
## EBOV HSA 23h
## paired-end, not strand specific, 100bp
#######################################

##PARAMETERS SHARED
PROJECT=ebov_hsa_23h
R1=ebov_hsa_23h_1.fastq
R2=ebov_hsa_23h_2.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=29
kmer3=33
kmer4=37
kmer5=41


###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink 25 --maxk 41 --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/test.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/test.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades/
rnaspades.py -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=4
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink 25 --maxk 41 --step 4
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
cd $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -o $DIR/binpacker
cd ..
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
###################################################################################




Assembly of Homo sapiens simulated (download script)


#######################################
## HSA FLUX SIMULATED, 2x100bp, not strand specific
#######################################

##PARAMETERS SHARED
PROJECT=hsa_flux
R1=hsa_flux_1.fastq
R2=hsa_flux_2.fastq
THREADS=48
MEM=400
DIR=~/$PROJECT
FINAL_DIR=~/$PROJECT/final

mkdir -p $DIR
mkdir -p $FINAL_DIR

kmer1=25
kmer2=35
kmer3=45
kmer4=55
kmer5=65


###################################################################################
## TRINITY
mkdir -p $DIR/trinity
Trinity --seqType fq --max_memory $MEM"G" --left $R1 --right $R2 --CPU $THREADS --output $DIR/trinity/
ln -s $DIR/trinity/Trinity.fasta $FINAL_DIR/trinity.fasta
###################################################################################

###################################################################################
## TRANS-ABYSS
mkdir -p $DIR/transabyss

for kmer in $kmer1 $kmer2 $kmer3 $kmer4 $kmer5; do
name=k${kmer}
assemblydir=$DIR/transabyss/${name}
mkdir -p $assemblydir
finalassembly=${assemblydir}/${name}-final.fa
transabyss -k ${kmer} --pe $R1 $R2 --outdir ${assemblydir} --name ${name} --threads $THREADS
done

mergedassembly=$DIR/transabyss/merged.fa
finalassembly1=$DIR/transabyss/k"$kmer1"/k"$kmer1"-final.fa
finalassembly2=$DIR/transabyss/k"$kmer2"/k"$kmer2"-final.fa
finalassembly3=$DIR/transabyss/k"$kmer3"/k"$kmer3"-final.fa
finalassembly4=$DIR/transabyss/k"$kmer4"/k"$kmer4"-final.fa
finalassembly5=$DIR/transabyss/k"$kmer5"/k"$kmer5"-final.fa
transabyss-merge --threads $THREADS --mink $kmer1 --maxk $kmer5 --prefixes k${kmer1}. k${kmer2}. k${kmer3}. k${kmer4}. k${kmer5}. --out ${mergedassembly} ${finalassembly1} ${finalassembly2} ${finalassembly3} ${finalassembly4} ${finalassembly5}
ln -s $DIR/transabyss/merged.fa $FINAL_DIR/trans-abyss.fasta
###################################################################################

###################################################################################
## SOAP-TRANS
mkdir -p $DIR/soap-trans
CONFIG=$DIR/soap-trans/$PROJECT.config
touch $CONFIG
echo '#maximal read length' > $CONFIG
echo "max_rd_len=100" >> $CONFIG
echo "[LIB]" >> $CONFIG
echo "#maximal read length in this lib" >> $CONFIG
echo "rd_len_cutoff=100" >> $CONFIG
#echo "#average insert size" >> $CONFIG
#echo "avg_ins=200" >> $CONFIG
echo "#if sequence needs to be reversed" >> $CONFIG
echo "reverse_seq=0" >> $CONFIG
echo "#in which part(s) the reads are used" >> $CONFIG
echo "asm_flags=3" >> $CONFIG
#echo "q=#{@reads}" >> $CONFIG #if @type == :SE
echo "#fastq paired end files" >> $CONFIG
echo "q1="$R1 >> $CONFIG
echo "q2="$R2 >> $CONFIG

kmer_min=$kmer1
kmer_max=$kmer5

# run soap with defualt settings
mkdir -p $DIR/soap-trans/default
SOAPdenovo-Trans-127mer all -s $DIR/soap-trans/$PROJECT.config -o $DIR/soap-trans/default/k23 -p $THREADS
ln -s $DIR/soap-trans/default/k23.scafSeq $FINAL_DIR/soap-trans-default.fasta
###################################################################################

###################################################################################
## RNA-SPADES
mkdir -p $DIR/rnaspades
rnaspades.py -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/rnaspades/
ln -s $DIR/rnaspades/transcripts.fasta $FINAL_DIR/rna-spades.fasta
###################################################################################

###################################################################################
## SPADES
mkdir -p $DIR/spades
spades.py --sc -1 $R1 -2 $R2 -t $THREADS -m $MEM --cov-cutoff auto -o $DIR/spades/
ln -s $DIR/spades/scaffolds.fasta $FINAL_DIR/spades.fasta
###################################################################################

###################################################################################
## OASES
k_min=$kmer1
k_max=$((kmer5 + 1))
k_step=10
type=-shortPaired

mkdir -p $DIR/oases
oases_pipeline.py -m $k_min -M $k_max -s $k_step -o $DIR/oases/ -d " $type -fastq $R1 $R2 -separate " -c
ln -s $DIR/oases/Merged/transcripts.fa $FINAL_DIR/oases.fasta
###################################################################################

###################################################################################
## IDBA-TRANS
###this tools assume the paired-end reads are in order (->, <-). If your data is in order (<-, ->), please convert it by yourself.
mkdir $DIR/idba-tran
fq2fa --merge --filter $R1 $R2 $DIR/idba-tran/r12.fasta
idba_tran -r $DIR/idba-tran/r12.fasta -o $DIR/idba-tran/ --mink $kmer1 --maxk $kmer5 --step 10 --num_threads $THREADS
ln -s $DIR/idba-tran/contig.fa $FINAL_DIR/idba-tran.fasta
###################################################################################

###################################################################################
## BRIDGER
OUT=$DIR/bridger/
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT
Bridger.pl --seqType fq --left $R1 --right $R2 --CPU $THREADS -o $OUT # start second time, because first run crashs at some point and then finishs
ln -s $OUT/Bridger.fasta $FINAL_DIR/bridger.fasta
###################################################################################

###################################################################################
## BINPACKER
OUT=$DIR/binpacker/
mkdir -p $OUT
BinPacker -s fq -p pair -l $R1 -r $R2 -o $OUT
ln -s $OUT/BinPacker_Out_Dir/BinPacker.fa $FINAL_DIR/binpacker.fasta
###################################################################################

###################################################################################
## SHANNON
mkdir $DIR/shannon
python shannon.py -p $THREADS -o $DIR/shannon --left $R1 --right $R2
ln -s $DIR/shannon/shannon.fasta $FINAL_DIR/shannon.fasta
###################################################################################

###################################################################################
## MIRA
mkdir $DIR/mira
touch $DIR/mira/manifest.config
echo "project = $PROJECT" > $DIR/mira/manifest.config
echo "job = est,denovo,accurate" >> $DIR/mira/manifest.config
echo "parameters = -DI:trt=/mnt/dessertlocal/projects/transcriptome_assembly/tmp -NW:cnfs=no -NW:cmrnl=no -SK:mmhr=1" >> $DIR/mira/manifest.config
# if @type == :SE
# config << "# defining illumina single end reads"
# config << "readgroup = se"
# config << "data = #{@reads}"
# config << "technology = #{@technology}"
# end
echo "# defining illumina paired end reads" >> $DIR/mira/manifest.config
echo "readgroup = pe" >> $DIR/mira/manifest.config
echo "data = $R1 $R2" >> $DIR/mira/manifest.config
echo "technology = solexa" >> $DIR/mira/manifest.config
echo "template_size = 50 1000 autorefine" >> $DIR/mira/manifest.config
echo "segment_placement = ---> <---" >> $DIR/mira/manifest.config

mira -c $DIR/mira -t $THREADS $DIR/mira/manifest.config
ln -s $DIR/mira/"$PROJECT"_assembly/"$PROJECT"_d_results/"$PROJECT"_out.unpadded.fasta $FINAL_DIR/mira.fasta
###################################################################################











S4: HISAT2 re-mapping rate

We mapped the quality-controlled reads back to their assembled contigs, to determine the amount of reads that were included in the assembly process by each tool.

S5: RNAQuast statistics

Overview of RNA-Quast statistical output. Please click on the corresponding link to get a short summary of the assembly comparisons or to observe the full output.

• Escherichia coli: short report (pdf) | short report (tsv) | full report + plots (html)
• Candida albicans: short report (pdf) | short report (tsv) | full report + plots (html)
• Arabidopsis thaliana: short report (pdf) | short report (tsv) | full report + plots (html)
• Mus musculus: short report (pdf) | short report (tsv) | full report + plots (html)
• Homo sapiens: short report (pdf) | short report (tsv) | full report + plots (html)
• Homo sapiens + EBOV 3h: short report (pdf) | short report (tsv) | full report + plots (html)
• Homo sapiens + EBOV 7h: short report (pdf) | short report (tsv) | full report + plots (html)
• Homo sapiens + EBOV 23h: short report (pdf) | short report (tsv) | full report + plots (html)
• Homo sapiens simulated: short report (pdf) | short report (tsv) | full report + plots (html)

S6: TransRate

TransRate: reference-free quality assessment of de novo transcriptome assemblies (27252236)

• TransRate results for Escherichia coli: download (CSV) | show (HTML)
• TransRate results for Candida albicans: download (CSV) | show (HTML)
• TransRate results for Arabidopsis thaliana: download (CSV) | show (HTML)
• TransRate results for Mus musculus: download (CSV) | show (HTML)
• TransRate results for Homo sapiens: download (CSV) | show (HTML)
• TransRate results for Homo sapiens + EBOV 3h: download (CSV) | show (HTML)
• TransRate results for Homo sapiens + EBOV 7h: download (CSV) | show (HTML)
• TransRate results for Homo sapiens + EBOV 23h: download (CSV) | show (HTML)
• TransRate results for Homo sapiens simulated: download (CSV) | show (HTML)

S7: ExN50

An alternative to the standard contig Nx (e.g. N50) statistic that can be more appropriate for transcriptome assembly is the ExN50 statistic. The N50 statistic is computed as usual but limited to the top most highly expressed transcripts that represent x% of the total normalized expression data. Therefore, we first calculated transcript abundances using the alignment-free quantification tool Salmon and subsequently used the Trinity utilities to calculate ExN50 values. We report the Ex90N50 metric as one of our main metrics.

S8: BUSCO

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BUSCO: Assessing genome assembly and annotation completeness with Benchmarking Universal Single-Copy Orthologs (26059717)

Escherichia coli	Candida albicans	Arabidopsis thaliana
Homo sapiens + EBOV 3h	Homo sapiens + EBOV 7h	Homo sapiens + EBOV 23h
Mus musculus	Homo sapiens	Homo sapiens simulated






S9: DETONATE

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DETONATE: DE novo TranscriptOme rNa-seq Assembly with or without the Truth Evaluation (25608678).

Escherichia coli DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-210712852.43	0.817076	0.0320276	0.639604
oases	-245161321.58	0.79302	0.0264751	0.487533
trans-abyss	-151118336.50	0.822888	0.0328714	0.611844
soap-trans-default	-340812817.07	0.861768	0.062212	0.735111
bridger	-174824953.19	0.811427	0.0303294	0.635494
binpacker	-262761906.88	0.162242	0.000152509	0.0646008
idba-tran	-437779699.97	0.828836	0.028401	0.661802
shannon	-197944222.32	0.807803	0.026662	0.62028
spades	-194007062.42	0.874921	0.0582104	0.711878
rna-spades	-200089085.55	0.843184	0.0697163	0.716887





Candida albicans DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-334561156.85	0.679298	0.0766464	0.726441
oases	-419101803.95	0.617874	0.0756063	0.505806
trans-abyss	-341314448.16	0.734363	0.0836716	0.594869
soap-trans-default	-448426551.63	0.603828	0.0627947	0.730318
bridger	-354594754.45	0.681131	0.0647035	0.742303
binpacker	-355541613.29	0.681339	0.0650714	0.725447
idba-tran	-611471453.61	0.552593	0.0463257	0.727045
shannon	-356256806.38	0.718923	0.0686033	0.635121
spades	-369270117.24	0.707372	0.0618753	0.744748
rna-spades	-352586681.38	0.691902	0.0555932	0.738039





Arabidopsis thaliana DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-458951076.16	0.706259	0.0503788	0.713888
oases	-601376841.14	0.657365	0.0315914	0.421727
trans-abyss	-452320279.03	0.742054	0.0417782	0.586087
soap-trans-default	-733935298.80	0.616916	0.0417285	0.766783
bridger	-540615846.28	0.664801	0.0413928	0.717523
binpacker	-963432760.48	0.479321	0.00483293	0.255829
idba-tran	-671741034.20	0.656141	0.0324962	0.762677
shannon	-1343654128.47	0.373612	0.0404999	0.631825
spades	-593763835.97	0.713745	0.0592855	0.792133
rna-spades	-573548801.65	0.720746	0.0674779	0.786267





Mus musculus DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-2226879012.19	0.663224	0.00924793	0.424159
oases	-3338075293.16	0.466246	0.00191568	0.185852
trans-abyss	-2152573788.62	0.671318	0.0245507	0.399779
soap-trans-default	-2638014207.45	0.55072	0.0232672	0.509524
bridger	-2376224869.25	0.601723	0.00662728	0.450365
binpacker	-4888140588.38	0.284626	8.78969e-05	0.0681366
idba-tran	-5029841952.79	0.531343	0.0107357	0.495647
shannon	-2938129166.97	0.559955	0.00746879	0.38078
spades	-3008040595.31	0.686892	0.0051975	0.522144
rna-spades	-2447339857.69	0.714442	0.0143569	0.498195





Homo sapiens DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-6510590491.36	0.511762	0.0161655	0.42599
oases	-11817309171.03	0.243335	0.0189894	0.181064
trans-abyss	-6255709753.29	0.550435	0.203218	0.509015
soap-trans-default	-9033548866.99	0.373036	0.205052	0.566304
bridger	-7718007001.60	0.398725	0.010466	0.480306
binpacker	-10026565572.34	0.366908	0.000374367	0.1451
idba-tran	-16304023086.75	0.286957	0.0168006	0.552302
shannon	-8959903268.75	0.420737	0.0229751	0.346262
spades	-12123853339.02	0.389103	0.0140352	0.606564
rna-spades	-7163901001.16	0.427956	0.0128943	0.619367





Homo sapiens + EBOV 3h DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-1235651692.60	0.518515	0.0149362	0.457458
oases	-1726915020.53	0.400049	0.0149624	0.213295
trans-abyss	-1196653162.51	0.541571	0.0469628	0.458425
soap-trans-default	-1348871374.03	0.452867	0.0387229	0.558308
bridger	-1245885780.74	0.507537	0.0134374	0.491961
binpacker	-2956358196.35	0.216398	6.61916e-05	0.0537161
idba-tran	-1840189936.95	0.435537	0.0135966	0.552636
shannon	-1264628787.04	0.516653	0.0279902	0.388568
spades	-1316600593.27	0.493276	0.0130479	0.571439
rna-spades	-1199874337.85	0.542847	0.022366	0.610776





Homo sapiens + EBOV 7h DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-1621797552.26	0.548502	0.0133497	0.461589
oases	-2354768025.10	0.446502	0.01377	0.204378
trans-abyss	-1644765267.08	0.565015	0.0474134	0.456358
soap-trans-default	-1853762729.85	0.489643	0.0382348	0.552421
bridger	-1803791019.01	0.509876	0.0115023	0.489854
binpacker	-1948162277.75	0.498755	0.000981932	0.303781
idba-tran	-2796730511.36	0.433572	0.0122321	0.555899
shannon	-1739716273.79	0.551822	0.0303068	0.373245
spades	-2194102523.92	0.481227	0.0139344	0.570306
rna-spades	-1712618020.99	0.555596	0.0208343	0.6074





Homo sapiens + EBOV 23h DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-1434423706.95	0.767084	0.0124217	0.44378
oases	-3869713310.16	0.20915	0.0121553	0.218292
trans-abyss	-1445566961.21	0.779996	0.044702	0.448004
soap-trans-default	-1599735160.94	0.694346	0.0354917	0.547701
bridger	-1543477872.71	0.739796	0.0109893	0.47975
binpacker	-2016262591.54	0.706858	0.00035399	0.203836
idba-tran	-4311460879.81	0.161691	0.0113647	0.551236
shannon	-4947278470.01	0.0955414	0.0282574	0.379485
spades	-3603623299.10	0.247863	0.0105064	0.564203
rna-spades	-1673532042.88	0.589338	0.0196731	0.601376





Homo sapiens simulated DETONATE results

	RSEM-EVAL score	KC score	Contig F1 score	Nucleotide F1 score
trinity	-2794698982.84	0.883827	0.0633606	0.566043
oases	-4615211068.41	0.738406	0.0488742	0.223824
trans-abyss	-2379646547.27	0.923633	0.0987237	0.679067
soap-trans-default	-5270177770.68	0.595837	0.0752428	0.733277
bridger	-3663922534.65	0.81708	0.04805	0.710854
binpacker	-3649658509.59	0.821483	0.0475004	0.601458
idba-tran	-7528195849.86	0.59977	0.0504635	0.783647
shannon	-11513711720.10	0.261483	0.0695831	0.453332
spades	-4779916710.32	0.773079	0.0528581	0.789225
rna-spades	-3911011816.48	0.785485	0.0541619	0.695881





S10: Selected main metrics

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Based on our experiences and in comparison to other studies (see manuscript for details) we selected 20 metrics to get an initial idea of the performance of each assembler on each data set. For each metric and assembly we calculated (0,1)-normalized scores displayed in subscript next to the statistical value.

Escherichia coli selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	77.010.7	3470.42	130.87	0.3680.88	467.0350.2	3960.95	1.0120.98	3690.46	4070.92	0.316351.0	75.03611.0	NA	NA	21430.43	0.6396040.86	0.03202760.46	0.8170760.92	-0.210.79	2840.83	1900.96
Oases	49.160.24	7431.0	1030.0	0.7060.7	457.7170.19	4141.0	1.4430.0	2290.21	4180.94	0.305580.97	65.717260.69	NA	NA	36020.0	0.4875330.63	0.02647510.38	0.793020.89	-0.250.66	2990.88	1721.0
Trans_ABySS	95.671.0	4460.56	180.83	0.3220.9	310.4420.0	320.0	1.0440.9	1770.12	4360.99	0.028440.07	72.730440.92	NA	NA	30810.15	0.6118440.82	0.03287140.47	0.8228880.93	-0.151.0	2970.88	1701.0
SOAPdenovo_Trans	56.620.36	4390.55	10.99	0.1341.0	364.8810.07	2610.6	1.0021.0	1730.11	4230.96	0.194920.61	71.988070.9	NA	NA	25790.3	0.7351111.0	0.0622120.89	0.8617680.98	-0.340.34	3160.94	1780.99
Bridger	87.350.86	3630.44	520.5	0.2650.93	472.8060.21	3820.92	1.0140.97	3930.5	4020.9	0.310820.98	74.590620.99	NA	NA	21170.44	0.6354940.85	0.03032940.43	0.8114270.91	-0.170.93	2850.83	1900.96
BinPacker	71.090.6	660.0	80.92	2.0510.0	1100.7741.0	340.01	1.2670.4	6751.0	370.0	0.006190.0	45.133380.0	NA	NA	1911.0	0.06460080.0	0.0001525090.0	0.1622420.0	-0.260.62	500.0	7110.0
IDBA_Tran	34.310.0	4140.51	01.0	0.1670.98	540.8280.29	2500.57	1.0011.0	3040.34	3960.89	0.160290.5	71.993010.9	NA	NA	20690.45	0.6618020.89	0.0284010.41	0.8288360.94	-0.440.0	2960.87	1960.95
Shannon	76.690.69	3720.45	540.48	0.1850.97	482.6590.22	3930.95	1.0270.94	3630.45	3990.9	0.30490.96	73.34470.94	NA	NA	21450.43	0.620280.83	0.0266620.38	0.8078030.91	-0.20.83	2800.82	1980.95
SPAdes_sc	88.040.88	4890.62	120.88	0.2480.94	558.9040.31	2100.47	1.0060.99	1110.0	4411.0	0.143910.44	69.469140.81	NA	NA	23700.36	0.7118780.97	0.05821040.83	0.8749211.0	-0.190.86	3321.0	1721.0
SPAdes_rna	88.90.89	3400.4	100.9	0.2120.96	359.1950.06	1870.41	1.0090.98	2450.24	3380.75	0.175870.55	73.776770.96	NA	NA	25560.31	0.7168870.97	0.06971631.0	0.8431840.96	-0.20.83	2550.73	1890.96

Escherichia coli summary of (0,1)-normalized scores

• Trinity: 13.61/18
• Oases: 10.36/18
• Trans_ABySS: 11.53/18
• SOAPdenovo_Trans: 12.59/18
• Bridger: 13.55/18
• BinPacker: 5.55/18
• IDBA_Tran: 11.49/18
• Shannon: 13.08/18
• SPAdes_sc: 13.37/18
• SPAdes_rna: 12.85/18

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Candida albicans selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	97.320.91	34170.09	720.89	1.8190.27	969.3270.56	41460.46	1.0390.97	18620.73	17320.76	0.183570.06	81.692610.51	0.455720.83	0.130150.85	93910.96	0.7264410.92	0.07664640.81	0.6792980.7	-0.331.0	13550.65	1640.7
Oases	93.610.6	98131.0	6130.0	2.1210.09	941.60.53	48080.79	2.250.0	15880.4	18360.91	0.235930.31	79.5370.33	0.058270.05	0.867920.0	260100.0	0.5058060.0	0.07560630.78	0.6178740.36	-0.420.68	13820.71	1400.79
Trans_ABySS	98.451.0	79790.74	4490.27	1.2880.58	773.5840.29	52141.0	1.9160.27	16640.49	18951.0	0.38241.0	81.468160.49	0.033630.0	0.821940.05	225810.2	0.5948690.37	0.08367161.0	0.7343631.0	-0.340.96	14610.87	1090.9
SOAPdenovo_Trans	95.120.73	27690.0	91.0	0.5691.0	567.1690.0	32900.03	1.0071.0	13740.14	12740.08	0.198890.13	84.146380.71	0.480330.88	0.018970.98	89470.98	0.7303180.94	0.06279470.44	0.6038280.28	-0.450.57	10420.0	2480.4
Bridger	96.720.86	40380.18	2360.62	1.9960.17	1131.3260.79	40050.39	1.1050.92	19230.8	16980.71	0.171960.0	79.399540.32	0.363080.65	0.312310.64	106510.88	0.7423030.99	0.06470350.49	0.6811310.71	-0.350.93	14160.78	1330.81
BinPacker	96.660.85	42670.21	2590.59	2.2830.0	1189.5520.87	40330.4	1.1360.89	18760.74	17010.71	0.171370.0	78.649890.26	0.367720.66	0.36050.59	110720.86	0.7254470.92	0.06507140.5	0.6813390.71	-0.360.89	14160.78	1350.81
IDBA_Tran	86.340.0	29490.03	81.0	0.8070.86	922.0960.5	32380.0	1.0031.0	12570.0	12230.0	0.19610.12	83.374130.65	0.414210.75	0.002891.0	86521.0	0.7270450.93	0.04632570.0	0.5525930.0	-0.610.0	10700.06	2400.43
Shannon	96.510.84	40770.19	3620.41	0.9990.75	678.9470.16	37360.25	1.4120.67	13210.08	14800.38	0.302640.62	87.689731.0	0.051880.04	0.484550.44	127930.76	0.6351210.54	0.06860330.6	0.7189230.92	-0.360.89	10880.1	3590.0
SPAdes_sc	97.510.92	35040.1	600.91	1.7990.28	1279.4111.0	44370.61	1.0011.0	20901.0	18470.93	0.174570.02	77.021730.13	0.542361.0	0.002941.0	91730.97	0.7447481.0	0.06187530.42	0.7073720.85	-0.370.86	15231.0	811.0
SPAdes_rna	97.430.92	39390.17	820.88	1.9590.19	1171.7140.85	45330.66	1.0590.95	20340.93	18370.91	0.198190.13	75.462750.0	0.406230.73	0.160620.82	105670.89	0.7380390.97	0.05559320.25	0.6919020.77	-0.350.93	14990.95	880.97

Candida albicans summary of (0,1)-normalized scores

• Trinity: 13.633/20
• Oases: 8.329/20
• Trans_ABySS: 12.486/20
• SOAPdenovo_Trans: 10.277/20
• Bridger: 12.639/20
• BinPacker: 12.243/20
• IDBA_Tran: 8.308/20
• Shannon: 9.632/20
• SPAdes_sc: 14.996/20
• SPAdes_rna: 13.86/20

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Arabidopsis thaliana selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	95.710.96	133570.33	12120.84	1.3150.79	978.2130.55	23020.8	1.1280.88	14510.83	72190.81	0.155120.69	74.790090.57	NA	NA	359640.61	0.7138880.85	0.05037880.73	0.7062590.9	-0.460.99	11171.0	2191.0
Oases	91.930.88	326471.0	68040.06	3.4710.42	1232.6880.89	27531.0	1.9760.0	13050.71	83611.0	0.192070.9	72.074970.38	NA	NA	787620.0	0.4217270.31	0.03159140.43	0.6573650.77	-0.60.83	11080.99	2480.97
Trans_ABySS	97.551.0	197140.55	72270.0	0.6120.91	745.0880.24	19600.65	1.3410.66	9400.41	81330.96	0.209921.0	72.504470.41	NA	NA	531800.37	0.5860870.62	0.04177820.59	0.7420541.0	-0.451.0	11191.0	2240.99
SOAPdenovo_Trans	85.310.73	87090.17	511.0	0.0991.0	561.7460.0	12130.32	1.0081.0	4490.0	54110.52	0.14790.65	80.777781.0	NA	NA	286320.72	0.7667830.95	0.04172850.59	0.6169160.66	-0.730.69	10580.93	2480.97
Bridger	90.640.85	128820.32	19950.73	2.4270.6	990.3490.57	19090.63	1.0670.94	16360.99	71630.8	0.1320.57	74.663130.56	NA	NA	330080.66	0.7175230.86	0.04139280.58	0.6648010.79	-0.540.9	11030.98	2290.99
BinPacker	67.150.34	37750.0	12330.84	5.9020.0	1312.1711.0	4890.0	1.1490.85	14520.83	22270.0	0.030290.0	66.836810.0	NA	NA	90811.0	0.2558290.0	0.004832930.0	0.4793210.29	-0.960.43	2620.0	11620.0
IDBA_Tran	89.040.82	95770.2	2010.98	0.1360.99	848.3560.38	11750.3	1.011.0	8050.3	60870.63	0.146130.64	80.707970.99	NA	NA	276660.73	0.7626770.95	0.03249620.44	0.6561410.77	-0.670.75	9300.78	2690.95
Shannon	51.530.0	120180.29	7690.9	0.7240.89	1044.9040.64	20040.67	1.1850.82	16541.0	65410.7	0.147770.65	76.083680.66	NA	NA	301780.7	0.6318250.7	0.04049990.57	0.3736120.0	-1.340.0	10490.92	2960.92
SPAdes_sc	94.290.93	111710.26	11890.84	0.430.94	944.3880.51	23180.81	1.0071.0	10300.48	78000.91	0.1510.67	74.471820.55	NA	NA	295690.71	0.7921331.0	0.05928550.87	0.7137450.92	-0.590.84	10740.95	2250.99
SPAdes_rna	93.670.92	111600.26	5050.94	0.3410.96	807.4220.33	22370.77	1.0270.98	12390.66	75010.86	0.154150.69	75.672170.63	NA	NA	306210.69	0.7862670.99	0.06747791.0	0.7207460.94	-0.570.87	10120.88	2640.95

Arabidopsis thaliana summary of (0,1)-normalized scores

• Trinity: 14.15/18
• Oases: 11.53/18
• Trans_ABySS: 12.36/18
• SOAPdenovo_Trans: 11.9/18
• Bridger: 13.31/18
• BinPacker: 5.57/18
• IDBA_Tran: 12.6/18
• Shannon: 11.03/18
• SPAdes_sc: 14.18/18
• SPAdes_rna: 14.3/18

Download Excel | csv | PDF | LaTeX



Mus musculus selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	92.140.89	218040.4	5640.99	0.840.85	1216.8630.52	60320.96	1.5470.39	4200.0	82100.97	0.218340.88	52.207670.82	0.221360.53	0.519730.43	787260.6	0.4241590.78	0.009247930.37	0.6632240.88	-2.230.97	38950.97	19590.98
Oases	89.290.82	518321.0	526680.0	0.6250.9	406.0840.0	1830.02	1.90.0	20440.48	62930.71	0.022110.09	45.357950.4	0.019210.0	0.909030.0	1889470.0	0.1858520.26	0.001915680.07	0.4662460.42	-3.340.59	30660.75	24760.85
Trans_ABySS	93.960.93	297190.56	6120.99	0.3160.97	752.8110.22	62811.0	1.750.17	5300.03	84011.0	0.247281.0	53.78860.92	0.115520.25	0.628360.31	1051530.46	0.3997790.73	0.02455071.0	0.6713180.9	-2.151.0	39921.0	19260.99
SOAPdenovo_Trans	90.980.86	126030.21	611.0	0.1631.0	519.9620.07	22440.35	1.0310.97	27960.7	73750.86	0.095390.38	51.454310.78	0.401381.0	0.119220.88	536110.74	0.5095240.97	0.02326720.95	0.550720.62	-2.640.83	36160.9	19870.97
Bridger	91.660.88	182660.33	26430.95	0.9130.83	1073.3950.43	22360.35	1.3480.62	31000.79	76390.9	0.099330.4	49.562060.66	0.180760.42	0.413640.55	669540.67	0.4503650.84	0.006627280.27	0.6017230.74	-2.380.92	39140.98	19260.99
BinPacker	54.310.0	20370.0	6280.99	4.6090.0	1972.8091.0	5540.08	1.3460.62	27150.68	10140.0	0.01540.06	38.855790.0	0.130360.29	0.600650.34	65141.0	0.06813660.0	8.78969e-050.0	0.2846260.0	-4.890.05	3100.0	58710.0
IDBA_Tran	70.640.38	122940.21	411.0	0.1870.99	811.990.26	9670.14	1.0051.0	16720.37	42080.43	0.091590.37	54.281470.95	0.294260.72	0.017111.0	455260.79	0.4956470.94	0.01073570.44	0.5313430.57	-5.030.0	26790.64	21880.92
Shannon	86.60.76	169150.3	19270.96	0.8020.86	877.7060.3	710.0	1.230.75	20670.49	52480.57	0.001070.0	55.043171.0	0.183940.43	0.426890.54	529480.75	0.380780.69	0.007468790.3	0.5599550.64	-2.940.73	28290.68	25340.84
SPAdes_sc	93.830.93	130990.22	5700.99	0.7060.88	1089.2640.44	26330.41	1.0031.0	38141.0	77360.91	0.084570.34	44.852870.37	0.379530.94	0.01341.0	515750.75	0.5221441.0	0.00519750.21	0.6868920.94	-3.010.7	38230.95	18771.0
SPAdes_rna	96.861.0	193670.35	10800.98	0.8020.86	910.4660.32	32690.51	1.1350.85	34180.88	78590.93	0.106240.43	43.578140.29	0.177180.41	0.332040.64	734610.63	0.4981950.95	0.01435690.58	0.7144421.0	-2.450.9	37730.94	18801.0

Mus musculus summary of (0,1)-normalized scores

• Trinity: 14.209/20
• Oases: 7.358/20
• Trans_ABySS: 14.429/20
• SOAPdenovo_Trans: 15.05/20
• Bridger: 13.501/20
• BinPacker: 5.102/20
• IDBA_Tran: 12.126/20
• Shannon: 11.576/20
• SPAdes_sc: 14.983/20
• SPAdes_rna: 14.46/20

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Homo sapiens selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	91.90.81	640610.22	33780.99	1.3820.74	795.2250.27	67880.99	2.3960.0	3260.0	89300.97	0.227440.87	50.815380.64	0.13180.3	0.590350.38	2864790.72	0.425990.59	0.01616550.08	0.5117620.87	-6.510.98	40040.96	18040.99
Oases	88.040.69	2074741.0	2161270.0	1.2530.77	343.4790.06	8680.1	2.3550.03	6660.17	80240.83	0.08810.33	42.088860.0	0.017930.0	0.940010.0	8432350.0	0.1810640.08	0.01898940.09	0.2433350.0	-11.820.45	35880.79	19220.93
Trans_ABySS	98.341.0	597790.2	27430.99	0.5730.93	246.8460.01	68241.0	1.7430.47	4410.06	91101.0	0.262191.0	51.922480.72	0.105880.23	0.632970.33	4378450.53	0.5090150.77	0.2032180.99	0.5504351.0	-6.261.0	41061.0	17701.0
SOAPdenovo_Trans	89.930.75	275290.03	2791.0	0.2691.0	218.00.0	22640.31	1.1870.87	7110.19	68060.64	0.091830.34	48.018340.44	0.267520.66	0.326750.67	2412360.78	0.5663040.89	0.2050521.0	0.3730360.42	-9.030.72	26250.39	21640.83
Bridger	86.830.66	432010.11	73290.97	1.4390.73	654.4130.21	21050.28	1.7080.5	13700.51	84400.89	0.0850.31	45.104190.22	0.140930.32	0.420730.57	2066350.83	0.4803060.71	0.0104660.05	0.3987250.51	-7.720.85	39090.92	18120.98
BinPacker	72.60.24	226110.0	56030.98	4.6290.0	2335.7291.0	28240.39	2.3890.01	23811.0	44560.26	0.073030.27	42.565740.04	0.069560.14	0.837540.11	729181.0	0.14510.0	0.0003743670.0	0.3669080.4	-10.030.62	20090.13	40780.0
IDBA_Tran	64.610.0	235160.0	3021.0	0.6660.91	487.1060.13	7090.07	1.0121.0	7080.19	27830.0	0.083260.31	52.459840.76	0.250390.61	0.022561.0	1386990.91	0.5523020.86	0.01680060.08	0.2869570.14	-16.30.0	16820.0	26150.63
Shannon	84.270.58	313280.05	28370.99	1.2620.77	711.8310.23	2420.0	1.530.63	13240.49	67580.63	0.003730.0	55.697821.0	0.066340.13	0.5030.48	1170680.94	0.3462620.42	0.02297510.11	0.4207370.58	-8.960.73	33850.7	21330.84
SPAdes_sc	92.040.81	310390.05	20220.99	0.8030.88	410.2240.09	17550.23	1.0151.0	11860.42	56760.46	0.080420.3	46.127570.3	0.396651.0	0.031190.99	1774770.86	0.6065640.97	0.01403520.07	0.3891030.47	-12.120.42	26250.39	22680.78
SPAdes_rna	95.950.93	498600.15	51260.98	1.2490.78	412.2380.09	32530.46	1.1920.87	7820.22	71550.69	0.111190.42	46.254250.31	0.232010.57	0.214490.79	2940830.71	0.6193671.0	0.01289430.06	0.4279560.6	-7.160.91	30890.58	19490.92

Homo sapiens summary of (0,1)-normalized scores

• Trinity: 12.375/20
• Oases: 6.307/20
• Trans_ABySS: 14.235/20
• SOAPdenovo_Trans: 11.92/20
• Bridger: 11.128/20
• BinPacker: 6.587/20
• IDBA_Tran: 8.611/20
• Shannon: 10.299/20
• SPAdes_sc: 11.475/20
• SPAdes_rna: 12.028/20

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Homo sapiens + EBOV 3h selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	90.730.94	271540.26	15800.94	1.7280.77	1083.5550.25	38260.97	1.6970.76	27530.6	81010.98	0.084340.67	54.506380.74	0.188690.3	0.59660.37	1094960.67	0.4574580.72	0.01493620.32	0.5185150.93	-1.240.98	39791.0	18291.0
Oases	86.920.88	981961.0	248450.0	2.3830.66	1308.2160.35	39481.0	3.910.0	24610.45	82881.0	0.123281.0	48.701840.32	0.021090.0	0.937240.0	3189360.0	0.2132950.29	0.01496240.32	0.4000490.56	-1.730.7	38030.95	18920.98
Trans_ABySS	93.91.0	307670.3	79560.68	0.7040.95	517.9160.02	31530.78	1.5540.81	19470.18	80720.97	0.116450.94	55.633720.82	0.11490.17	0.580880.38	1274480.61	0.4584250.73	0.04696281.0	0.5415711.0	-1.21.0	39821.0	18380.99
SOAPdenovo_Trans	89.80.93	131100.12	1481.0	0.4291.0	464.3680.0	25530.62	1.0420.99	23720.4	72190.86	0.071330.57	52.668490.6	0.358720.61	0.110180.89	627530.82	0.5583080.91	0.03872290.82	0.4528670.72	-1.350.91	35280.88	19340.97
Bridger	89.950.93	204340.19	18530.93	1.430.83	925.6190.19	27810.68	1.4340.85	26760.56	75190.9	0.070580.56	51.845730.54	0.199280.32	0.476790.5	831870.76	0.4919610.79	0.01343740.29	0.5075370.89	-1.250.97	38790.97	18620.99
BinPacker	36.60.0	16460.0	2790.99	6.1840.0	2899.3921.0	2550.0	2.0140.65	35301.0	4960.0	0.003610.0	44.282060.0	0.047230.05	0.831110.11	67521.0	0.05371610.0	6.61916e-050.0	0.2163980.0	-2.960.0	2050.0	59760.0
IDBA_Tran	78.410.73	135080.12	1781.0	0.7990.94	718.4020.11	5690.09	1.0051.0	19240.17	31210.34	0.06920.55	54.468860.73	0.426770.73	0.009061.0	544940.85	0.5526360.9	0.01359660.29	0.4355370.67	-1.840.64	21620.52	22160.9
Shannon	90.230.94	282480.28	21610.92	1.3130.85	921.0460.19	23030.55	1.8080.72	22360.33	70400.84	0.110890.9	58.165311.0	0.050620.05	0.693150.26	1041260.69	0.3885680.6	0.02799020.6	0.5166530.92	-1.260.97	35750.89	20960.93
SPAdes_sc	91.410.96	127750.12	12620.95	0.9440.91	661.3220.08	26530.65	1.0031.0	27350.59	75370.9	0.077290.62	51.507640.52	0.574751.0	0.0111.0	609000.83	0.5714390.93	0.01304790.28	0.4932760.85	-1.320.93	38900.98	18131.0
SPAdes_rna	93.831.0	171950.16	9430.97	1.0010.9	462.0870.0	33350.83	1.1170.96	16030.0	74100.89	0.084450.68	50.270420.43	0.352520.6	0.161380.84	942740.72	0.6107761.0	0.0223660.48	0.5428471.0	-1.21.0	36550.91	18510.99

Homo sapiens + EBOV 3h summary of (0,1)-normalized scores

• Trinity: 14.173/20
• Oases: 10.449/20
• Trans_ABySS: 14.35/20
• SOAPdenovo_Trans: 14.63/20
• Bridger: 13.64/20
• BinPacker: 4.808/20
• IDBA_Tran: 12.255/20
• Shannon: 13.418/20
• SPAdes_sc: 15.082/20
• SPAdes_rna: 14.351/20

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Homo sapiens + EBOV 7h selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	92.580.91	313190.16	12940.94	1.7460.6	1040.8750.29	46960.9	1.7570.78	28390.56	84580.93	0.082960.37	52.152560.77	0.190230.35	0.58220.39	1312840.82	0.4615890.64	0.01334970.27	0.5485020.87	-1.621.0	41191.0	17591.0
Oases	85.780.53	1155961.0	188930.0	2.5150.36	1364.3770.44	51151.0	4.3470.0	30030.64	88611.0	0.121490.9	43.815020.0	0.015720.0	0.947070.0	4134180.0	0.2043780.0	0.013770.28	0.4465020.1	-2.350.38	40550.97	18170.91
Trans_ABySS	94.221.0	374100.22	47690.75	0.7530.9	534.4020.04	40040.74	1.6630.8	20390.19	84050.92	0.128511.0	51.966260.75	0.072460.12	0.615660.35	1590920.74	0.4563580.63	0.04741341.0	0.5650151.0	-1.640.98	41211.0	17710.98
SOAPdenovo_Trans	90.520.79	152510.0	1251.0	0.4431.0	455.9590.0	32250.56	1.060.98	23850.35	75690.77	0.075710.27	49.378910.51	0.33140.64	0.134790.87	771710.97	0.5524210.86	0.03823480.8	0.4896430.43	-1.850.81	36670.77	18550.85
Bridger	89.140.72	234870.08	25090.87	1.4870.68	866.6080.2	29650.5	1.4510.87	27450.52	75400.77	0.075210.26	47.639590.35	0.177060.33	0.469340.51	1017880.9	0.4898540.71	0.01150230.23	0.5098760.58	-1.80.85	39790.93	17850.96
BinPacker	84.630.46	229280.08	26490.87	3.6740.0	2505.4261.0	28510.48	1.8840.74	37641.0	68440.64	0.056030.0	47.66020.35	0.113780.2	0.758620.2	822390.96	0.3037810.25	0.0009819320.0	0.4987550.5	-1.950.72	35900.73	23920.0
IDBA_Tran	76.390.0	161810.01	2031.0	0.8340.88	697.7480.12	7800.0	1.0071.0	19670.15	31970.0	0.075330.27	50.414160.61	0.389710.76	0.011161.0	675641.0	0.5558990.87	0.01223210.24	0.4335720.0	-2.80.0	21340.0	22170.28
Shannon	90.990.82	357850.2	26050.87	1.4260.7	918.8540.23	29640.5	1.9540.72	23280.32	75240.76	0.12250.92	54.68871.0	0.036240.04	0.709650.25	1341080.81	0.3732450.42	0.03030680.63	0.5518220.9	-1.740.9	37770.83	19550.69
SPAdes_sc	91.940.87	155980.0	20420.9	0.720.91	489.20.02	27950.46	1.0131.0	27660.53	70040.67	0.076180.28	48.010430.39	0.508021.0	0.025290.98	766460.97	0.5703060.91	0.01393440.28	0.4812270.36	-2.190.52	36440.76	18350.88
SPAdes_rna	93.40.95	218670.07	12190.94	1.0670.81	470.1610.01	38500.71	1.1610.95	16470.0	72260.71	0.090820.48	45.993420.2	0.298630.57	0.200120.8	1205970.85	0.60741.0	0.02083430.43	0.5555960.93	-1.710.92	35420.71	17800.97

Homo sapiens + EBOV 7h summary of (0,1)-normalized scores

• Trinity: 13.535/20
• Oases: 8.503/20
• Trans_ABySS: 14.111/20
• SOAPdenovo_Trans: 13.243/20
• Bridger: 11.81/20
• BinPacker: 9.171/20
• IDBA_Tran: 8.177/20
• Shannon: 12.505/20
• SPAdes_sc: 12.696/20
• SPAdes_rna: 13.004/20

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Homo sapiens + EBOV 23h selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	94.390.98	277850.23	10660.91	1.6840.63	1143.4920.26	40650.93	1.8060.73	29580.55	79351.0	0.073240.51	53.733730.46	0.140250.3	0.606690.36	1155780.72	0.443780.6	0.01242170.27	0.7670840.98	-1.431.0	38850.99	19441.0
Oases	70.050.6	835301.0	111940.0	2.3110.44	1500.7760.39	43301.0	3.970.0	31150.62	28680.0	0.107890.89	47.19380.02	0.031910.0	0.938110.0	3017800.0	0.2182920.04	0.01215530.27	0.209150.17	-3.870.31	38320.97	20140.97
Trans_ABySS	95.491.0	328660.3	32530.72	0.7690.91	578.8810.04	35750.79	1.6440.78	21780.21	78720.99	0.117471.0	53.731220.46	0.051680.05	0.614980.35	1317120.66	0.4480040.61	0.0447021.0	0.7799961.0	-1.450.99	39071.0	19541.0
SOAPdenovo_Trans	93.10.96	134180.04	941.0	0.4681.0	482.2570.0	27490.56	1.0490.98	24890.34	71220.84	0.068690.46	51.416920.3	0.395981.0	0.126490.88	645470.92	0.5477010.86	0.03549170.79	0.6943460.87	-1.60.95	34350.79	20420.96
Bridger	92.430.95	205600.13	18440.84	1.430.71	929.8570.17	27360.56	1.4590.85	28410.5	72220.86	0.068840.47	48.883160.13	0.196350.45	0.482010.49	876210.83	0.479750.69	0.01098930.24	0.7397960.94	-1.540.97	37840.94	19780.99
BinPacker	81.570.78	108620.0	14360.88	3.7550.0	3072.8981.0	15230.22	2.090.63	39651.0	32620.08	0.026310.0	46.967480.0	0.06750.1	0.816430.13	439131.0	0.2038360.0	0.000353990.0	0.7068580.89	-2.020.83	16950.0	43920.0
IDBA_Tran	48.370.27	141960.05	1341.0	0.8660.88	725.580.1	7140.0	1.0061.0	20480.15	31260.05	0.069540.47	52.334530.37	0.244310.58	0.010961.0	572710.95	0.5512360.87	0.01136470.25	0.1616910.1	-4.310.18	20420.16	24190.81
Shannon	31.290.0	150070.06	3810.97	0.5830.97	703.7480.09	16310.25	1.4350.85	17140.0	53490.49	0.077190.56	61.577021.0	0.048470.05	0.465020.51	608170.93	0.3794850.44	0.02825740.63	0.09554140.0	-4.950.0	28520.52	27640.67
SPAdes_sc	91.150.93	132450.03	12060.9	0.9510.85	649.4190.07	26540.54	1.0041.0	30710.6	71010.84	0.068370.46	49.530840.18	0.389490.98	0.018770.99	638720.92	0.5642030.91	0.01050640.23	0.2478630.22	-3.60.38	37100.91	19521.0
SPAdes_rna	93.120.96	186290.11	9080.93	1.150.79	476.1760.0	35350.78	1.1470.95	22030.22	69600.81	0.081930.61	47.538830.04	0.217370.51	0.192250.8	1020950.77	0.6013761.0	0.01967310.44	0.5893380.72	-1.670.93	34050.77	19920.98

Homo sapiens + EBOV 23h summary of (0,1)-normalized scores

• Trinity: 13.426/20
• Oases: 7.683/20
• Trans_ABySS: 13.865/20
• SOAPdenovo_Trans: 14.527/20
• Bridger: 12.722/20
• BinPacker: 7.552/20
• IDBA_Tran: 9.219/20
• Shannon: 8.989/20
• SPAdes_sc: 12.942/20
• SPAdes_rna: 13.126/20

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Homo sapiens simulated selected metrics

	Overall mapping rate	Transcripts >=1000nt	Misassemblies	Mismatches per transcript	Average alignment length	95%-assembled isoforms	Duplication ratio	Ex90N50	# full-length transcripts	Reference coverage	Mean ORF percentage	Optimal score	Percentage bases uncovered	Number of ambiguous bases	Nucleotide F1	Contig F1	KC score	RSEM EVAL	Complete BUSCOs	Missing BUSCOs
Trinity	96.390.95	84510.24	1390.97	1.2310.44	2261.3321.0	28980.96	2.1540.7	32340.96	14920.85	0.243960.34	43.378410.63	0.111680.22	0.502360.4	309900.77	0.5660430.61	0.06336060.31	0.8838270.94	-2.790.96	5880.94	260.99
Oases	73.260.62	281431.0	40940.0	2.1050.0	2090.550.88	29951.0	4.8540.0	33000.99	16721.0	0.375480.74	37.427360.1	0.01070.0	0.829810.0	1107230.0	0.2238240.0	0.04887420.03	0.7384060.72	-4.620.75	6131.0	221.0
Trans_ABySS	99.561.0	67030.17	1170.97	0.4590.82	939.3120.06	28770.95	1.5720.85	29020.78	13230.71	0.459441.0	45.491520.81	0.224310.47	0.316680.63	242820.84	0.6790670.81	0.09872371.0	0.9236331.0	-2.381.0	5370.8	211.0
SOAPdenovo_Trans	91.680.89	42630.07	490.99	0.3080.89	1009.080.11	16790.46	1.2740.93	28360.75	10060.45	0.182470.16	36.277480.0	0.205410.43	0.167580.81	152590.93	0.7332770.9	0.07524280.54	0.5958370.5	-5.270.68	2890.16	1090.74
Bridger	94.020.92	51670.11	5330.87	1.6520.23	1581.2990.51	15700.41	1.5070.87	32520.97	12100.62	0.18770.17	42.219870.52	0.142180.29	0.221250.74	180250.9	0.7108540.86	0.048050.01	0.817080.84	-3.660.86	5250.77	280.98
BinPacker	93.030.91	74240.2	7850.81	1.8490.13	1755.1550.64	16600.45	1.9490.76	29090.79	12460.65	0.202810.22	43.570580.64	0.129430.26	0.431880.49	258510.82	0.6014580.67	0.04750040.0	0.8214830.85	-3.650.86	5270.78	290.98
IDBA_Tran	85.340.79	27400.02	81.0	0.0961.0	859.9660.0	5770.0	1.0131.0	13970.0	4500.0	0.177640.14	47.62021.0	0.347940.74	0.012921.0	104890.97	0.7836470.99	0.05046350.06	0.599770.51	-7.530.44	2260.0	1420.65
Shannon	30.770.0	23410.0	660.99	0.3940.85	1061.0130.14	10560.2	1.4370.89	23680.51	6660.18	0.130610.0	47.367290.98	0.056170.1	0.174490.8	78011.0	0.4533320.41	0.06958310.43	0.2614830.0	-11.510.0	2420.04	3640.0
SPAdes_sc	97.020.96	26230.01	500.99	0.2150.94	979.9810.09	16250.43	1.0121.0	33151.0	10070.46	0.160270.09	39.618070.29	0.466921.0	0.01221.0	106740.97	0.7892251.0	0.05285810.1	0.7730790.77	-4.780.74	4070.47	550.9
SPAdes_rna	96.720.96	60530.14	3510.92	1.820.14	1649.1470.56	22200.68	1.5350.86	25490.6	11410.57	0.24590.35	40.382840.36	0.191160.4	0.247670.71	203020.88	0.6958810.83	0.05416190.13	0.7854850.79	-3.910.83	4620.61	260.99

Homo sapiens simulated summary of (0,1)-normalized scores

• Trinity: 14.165/20
• Oases: 9.833/20
• Trans_ABySS: 15.68/20
• SOAPdenovo_Trans: 11.403/20
• Bridger: 12.466/20
• BinPacker: 11.885/20
• IDBA_Tran: 10.306/20
• Shannon: 7.509/20
• SPAdes_sc: 13.219/20
• SPAdes_rna: 12.316/20

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S11: Runtime and memory consumption

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Shown are the runtime and max memory peak for each assembler and data set, run with 48 threads. For Trinity, we observed high memory peaks in the first few minutes of assembly. Here, we removed this high maximum memory peaks from the plots to show a better comparison of the memory usage over time.

S12: (0,1)-normalized scores per data set and metric

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For each metric and data set, we calculated normalized scores in the range of 0 and 1 (see manuscript methods) to compare the performance of the different assembly tools regarding this metric. For example, consider the E. coli data set and the metric Overall mapping rate that results in the following vector:

(77.01, 49.16, 95.67, 56.62, 87.35, 71.09, 34.31, 76.69, 88.04, 88.9)

The calculated (0,1)-normalized scores are:

(0.7, 0.24, 1.0, 0.36, 0.86, 0.6, 0.0, 0.69, 0.88, 0.89)

With this transformation, we aim to achive a somewhat fair comparison of the different assembly tools and metrics.

For each data set and assembly tool the scores are summed up to achieve a final score for comparison. Here we show a heat map for each data set representing the different normalized scores calculated for each metric and assembly tool. We used this heat maps to identify potentially highly correlated metrics.

• Escherichia coli normalized score heat map (PDF)
• Candida albicans normalized score heat map (PDF)
• Arabidopsis thaliana normalized score heat map (PDF)
• Mus musculus normalized score heat map (PDF)
• Homo sapiens normalized score heat map (PDF)
• Homo sapiens + EBOV 3h normalized score heat map (PDF)
• Homo sapiens + EBOV 7h normalized score heat map (PDF)
• Homo sapiens + EBOV 23h normalized score heat map (PDF)
• Homo sapiens simulated normalized score heat map (PDF)