Abyss-pe fungal genome assembly

Question

[Igor Lalin]

Would you please be able to take a look?

I am trying to assemble a fungal genome (40Mbp size) using abyss-pe at k=96. As recommended, after importing MiSeq DNA reads, first I concatanated two sets of reads, then trimmed using BWA style trimming at q=30, then used Spades read error correction tool after which finally I did the following: 1. abyss-mergepairs (error corrected Spades reads) to obtain reads that are overlapping 2. konnector (leftover abyss-mergepairs paired reads 1 and 2) to obtain reads that are non-overlapping 3. abyss-pe with paired-end leftovers from konnector, se reads from abyss-mergepais (overlapping reads) and se reads from konnector (non-overlapping reads) with using a long function to help with the scaffold

So this is the code:

cat fungusA_read1.fastq fungusB_read1.fastq > conc_read1.fastq
cat fungusA_read2.fastq fungusB_read2.fastq > conc_read2.fastq

trimBWAstyle -q30 conc_read1.fastq > trim_conc_read1.fastq
trimBWAstyle -q30 conc_read2.fastq > trim_conc_read2.fastq

spades --only-error-correction --careful -o spades_out -t 40 -k 96 \
-1 trim_conc_read1.fastq -2 trim_conc_read2.fastq

abyss-mergepairs -q30 spades_out/corrected/trim_conc_read1.00.0_0.cor.fastq \
spades_out/corrected/trim_conc_read2.00.0_0.cor.fastq

konnector -j 40 -k 96 -o kon96 out_reads1.fastq out_reads2.fastq

abyss-pe j=40 k=96 name=fungus lib='pe1 pe2' long='longa' \
                            pe1='../kon96_reads_1.fq' pe2='../kon96_reads_2.fq' \
                            se='../out_merged.fastq ../kon96_merged.fa' \
                            longa='../ref_genome.fa'

The problem is that abyss-pe hangs up after unitigs formation in the abyss-pe (last step). This is the last step nohup message:

Building the suffix array... Building the Burrows-Wheeler transform... Building the character occurrence table... Mateless 667892 100% Unaligned 0 Singleton 0 FR 0 RF 0 FF 0 Different 0 Total 667892 abyss-fixmate: error: All reads are mateless. This can happen when first and second read IDs do not match. error: pe1-3.hist': No such file or directory make: *** [pe1-3.dist] Error 1 make: *** Deleting filepe1-3.dist'

[lieven.sterck]

Most likely the issue is what it tells it is: the IDs from your forward and reverse read file do not correspond (or, the files are out of sync?).

All the other message are due to the fact you will end up with empty files afterwards.

Check you input fastq files (especially the PE ones) and double check if the IDs are correct.