No paired FASTA for Single-cell k-mer analysis

[ZhangDengwei]

Hi there,

Thanks for developing this amazing tool! I tried to apply it to paired-end single-cell sequencing reads, following the tutorial with the commands below

Rscript /data3/zhangdw/01.Toolkit/SAHMI/functions/run_kraken.r --sample SRR8088290 --fq1 ../../../02.FASTQ/scRNA_GSE121638/SRR8088290/SRR8088290_S1_L001_R1_001.fastq --fq2 ../../../02.FASTQ/scRNA_GSE121638/SRR8088290/SRR8088290_S1_L001_R2_001.fastq --out_path out. --Kraken2Uniq_path /data/zhangdw/01.Toolkit/anaconda3/envs/py37/bin/kraken2 --kraken_database_path /data3/zhangdw/02.Database/01.Genome/10.Kranken2/03.Default_plus_human/kdb_human/ --kreport2mpa_path /data3/zhangdw/01.Toolkit/SAHMI/functions/kreport2mpa.py --paired T
Rscript /data3/zhangdw/01.Toolkit/SAHMI/functions/extract_microbiome_reads.r --sample_name SRR8088290 --fq out.SRR8088290_1.fq --fq out.SRR8088290_2.fq --kraken_report out.SRR8088290.kraken.report.txt --mpa_report out.SRR8088290.kraken.report.mpa.txt --out_path ./
Rscript /data3/zhangdw/01.Toolkit/SAHMI/functions/extract_microbiome_output.r --sample_name SRR8088290 --output_file out.SRR8088290.kraken.output.txt --kraken_report out.SRR8088290.kraken.report.txt --mpa_report out.SRR8088290.kraken.report.mpa.txt --out_path ./ 

Prior to running k-mer analysis, generated files include

-rw-rw-r--. 1 dwzhang dwzhang    736720185 Oct 16 20:03 SRR8088290.fa
-rw-rw-r--. 1 dwzhang dwzhang    506639410 Oct 16 20:10 SRR8088290.microbiome.output.txt
-rw-rw-r--. 1 dwzhang dwzhang  39468201318 Oct 16 19:35 out.SRR8088290.kraken.output.txt
-rw-rw-r--. 1 dwzhang dwzhang      1747721 Oct 16 19:35 out.SRR8088290.kraken.report.mpa.txt
-rw-rw-r--. 1 dwzhang dwzhang       618413 Oct 16 19:35 out.SRR8088290.kraken.report.std.txt
-rw-rw-r--. 1 dwzhang dwzhang       695614 Oct 16 19:35 out.SRR8088290.kraken.report.txt
-rw-rw-r--. 1 dwzhang dwzhang 132120472618 Oct 16 19:35 out.SRR8088290_1.fq
-rw-rw-r--. 1 dwzhang dwzhang 132120472618 Oct 16 19:35 out.SRR8088290_2.fq

I wonder why only one SRR8088290.fa was generated from the paired reads, which only include reads from out.SRR8088290_2.fq but not out.SRR8088290_1.fq. Also, sckmer.r can take in two paired fasta files. Is there anything wrong regarding my codes or my understanding of the tutorial?

Any suggestion would be greatly appreciated!

[liyongzheng1]

I also encountered the same problem, how did you solve it? @ZhangDengwei

[ZhangDengwei]

Run individually for read 1 and read 2.

Rscript extract_microbiome_reads.r --sample_name ${q}_1 --fq out.${q}_1.fq --kraken_report out.${q}.kraken.report.txt --mpa_report out.${q}.kraken.report.mpa.txt --out_path ./
Rscript extract_microbiome_reads.r --sample_name ${q}_2 --fq out.${q}_2.fq --kraken_report out.${q}.kraken.report.txt --mpa_report out.${q}.kraken.report.mpa.txt --out_path ./

[liyongzheng1]

OK, I misunderstood that I need to repeat the -fq parameter twice, Thank you!