Dear Biostars, Hi
I have the RNA-seq data of a salmon fish (3 cond1 and 3 cond2 as biological replicate and both are paired-ends -> 12 fastq files ) and I have done Trinity de novo assembly and DEG analysis on them.
Recently the draft genome of that salmon species have been released.
I want to run a genome-guided and then DEG analysis on the same data to compare the results.
Using many helps from Biostars and @Kevin Blighe, I have selected HISAT2, StringTie approach for this purpose.
First, I created an indexed genome using: ./hisat2-build -p 6 '/home/Salmon-genome/GCF_salmon_genome.fna' ht2_base_salmon_genome
and now I have 8 *.ht2 files and want to map my reads to that reference.
Q1: Please check below mapping script I have found from here to see if it is correct for my case or not. specially I need help about paired-end data mapping. what should I write instead of sample_1.fastq in this case that I have paired-end reads?
./hisat2 -p 6 -x path/to/reference/fileName -1 path/to/fastqFile/sample_1.fastq -2 path/to/fastqFile/sample_2.fastq -S /path/to/outDir/fileName.sam &> /path/to/outDir/fileName.sam.info
Q2: can I map all my 12 files to the reference genome in one script? how? (is it by using && and repeating the script, changing fastq file names?).
Thanks
Q1: Sample_1.fastq will be replaced by R1 data file. Sample_2.fastq will be replaced by R2 data file.
Q2: You could follow parallel approach (Script to run blast locally with multiple files in a directory as queries ) or use this method Hisat2 multiple paired end reads
Dear @genomax, Hi and thank you. I guess all other parameters in the script were fine.
would you please explain more what do you mean by "R1 data file" ?
My fastq files names are as :
J1-left.fq, J1-right.fq, J2-left.fq, J2-right.fq, J3-left.fq, J3-right.fq, H1-left.fq, H1-right.fq, H2-left.fq, H2-right.fq, H3-left.fq and H3-right.fq.
Hi there, I have used "./hisat2 -p 6 -x ht2_base_salmon_genome -1 '/media/Seagate Backup Plus Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_left.fq' -2 '/media/Seagate Backup Plus Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_right.fq' -S H1.sam &> H1.sam.info"
and then I received this warning in the H1.sam.info file:
Warning: Same mate file "/media/Seagate" appears as argument to both -1 and -2
Extra parameter(s) specified: "Backup", "Plus", "Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_left.fq",
"Backup", "Plus", "Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_right.fq" Note that if <mates> files are specified using -1/-2, a <singles> file cannot also be specified. Please run bowtie separately for mates and singles.
Error: Encountered internal HISAT2 exception (#1) Command:
/home/Desktop/salmon-genome-2018/hisat2-2.1.0/hisat2-align-s --wrapper basic-0 -p 6 -x ht2_base_salmon_genome -S H1.sam -1 /media/Seagate -2 /media/Seagate Backup Plus Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_left.fq Backup Plus Drive/Bioinformatics/RNA-Seq/RNA_Seq_Data/clean_reads/H1/H1_clean_right.fq (ERR): hisat2-align exited with value 1
See if this helps: https://askubuntu.com/questions/731579/problem-in-renaming-the-partition-and-opening-a-file-through-terminal
If your drive is formatted using ntfs (windows file system) then are you able to write to it under ubuntu? As I recall that requires installation of some additional software (which you may have done).
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