PBcR pipeline(S) was proposed in the ref (Reducing assembly complexity of microbial genomes with single-molecule sequencin, Genome Biology 2013).

PBcR pipeline

Following we used E. coli as an example to show the steps. All the necessary programs were downloaded from cbcb (or direct download the package) and more detail information was described at PacBioToCA

1. Long reads self-correction

pacBioToCA -length 500 -partitions 200 -l pacbio -t 6 -s pacbio.spec -fastq Filtered_four.fastq longReads=1 genomeSize=4650000

2. Trimming the corrected long reads

python trimFastqByQVWindow.py --qvCut=54.5 --out=trimmed.pacbio.fastq pacbio.fastq

3. Convert the data format from fastq to frg

java convertFastqToFastaAndQual trimmed.pacbio.fastq trimmed.pacbio.fasta trimmed.pacbio.qual
convert-fasta-to-v2.pl -l Pacbio -s trimmed.pacbio.fasta -q trimmed.pacbio.qual > trimmed.pacbio.frg

4. Select 25X longest corrected long reads

gatekeeper -T -F -o asm.gkpStore trimmed.pacbio.frg
gatekeeper -dumpfasta 25X_Clr -longestlength 0 116250000 asm.gkpStore
gatekeeper -dumpfrg -longestlength 0 116250000 asm.gkpStore > 25X_Clr.frg

5. Assemble

runCA -p asm -d asm -s asm.spec 25X_Clr.frg

Dataset 5 (E. coli K-12 MG1655, 17 SMRT cells)

We used all SMRT cells and randomly selected four, six and eight SMRT cells three times for each, and evaluated the assemblies by QUAST against the reference genome (NC_000913) and Ec_gene_list. (more detail)

Performance

Dataset 6 (E.coli K-12 MG1655, 8 SMRT cells)

We used all SMRT cells and randomly selected four and six SMRT cells three times for each, and evaluated the assemblies by QUAST against the reference genome (NC_000913) and Ec_gene_list. (more detail)

Performance

Misassemblies for Adobe reader.

Dataset 7, (M. ruber DSM1279, 4 SMRT cells)

We used all SMRT cells to do assembly and evaluated the assemblies by QUAST against the reference genome (NC_013946) and Mr_gene_list. (more detail)

Performance

Dataset 8 (P. heparinus DSM1279, 7 SMRT cells)

We used all SMRT cells and randomly selected four SMRT cells three times for each, and evaluated the assemblies by QUAST against the reference genome (NC_013061) and Ph_gene_list. (more detail)

Performance

Misassemblies for Adobe reader.

Dataset 9 (E. coli K-12, P4-C2 chemistry, 20 Kbp, 1 SMRT cell)

We used all SMRT cells and evaluated the assemblies by QUAST against the reference genome (NC_000913) and Ec_gene_list. (more detail)

Performance