How many read files I can use in tophat in the same run?

Question

[zizigolu]

Good morning friends,

Supposing I have and experiment with 10 samples and each with just sra file. In tophat -r 20 test_ref reads_1.fq reads_2.fq syntax I should mention all of my fastq files? If yes, may you please tell me the order of files. I mean for example I should type reads_1.fq -> reads_10.fq?

Thanks a lot

[Anushka]

Run tophat on each sample separately (or write a simple bash script with a list of fastq file names as input and specify different output directory for each of them). In case of paired-end reads_1.fq and reads_2.fq defines for Right and Left.

Answer 1

[Devon Ryan]

Each sample must be aligned independently. You can not give tophat all of the files for all of the samples at one time and get multiple output files.

[zizigolu]

Thanks Devon,

Sorry

I placed the orf_coding.fasta (my reference genome) in the bowtie2 file and indexed the genome in the same bowtie2, and run this syntax but error,

May you please tell me a solution?

[izadi@lbox161 bowtie2-2.2.5]$ $TOP/tophat -r 20 orf_coding.fasta SRR1944914_trimmed.fastq SRR1944926_trimmed.fastq

[2015-09-01 10:27:50] Beginning TopHat run (v2.1.0)
-----------------------------------------------
[2015-09-01 10:27:50] Checking for Bowtie
          Bowtie version:     2.2.5.0
[2015-09-01 10:27:50] Checking for Bowtie index files (genome)..
Error: Could not find Bowtie 2 index files (orf_coding.fasta.*.bt2)
[izadi@lbox161 bowtie2-2.2.5]$ $TOP/tophat -r 20 orf_coding.fasta SRR1944914_trimmed_unmapped.fastq SRR1944926_trimmed_unmapped.fastq

[2015-09-01 10:29:01] Beginning TopHat run (v2.1.0)
-----------------------------------------------
[2015-09-01 10:29:01] Checking for Bowtie
          Bowtie version:     2.2.5.0
[2015-09-01 10:29:01] Checking for Bowtie index files (genome)..
Error: Could not find Bowtie 2 index files (orf_coding.fasta.*.bt2)
[izadi@lbox161 bowtie2-2.2.5]$

[Devon Ryan]

This part of the error message should tell you what to fix:

Error: Could not find Bowtie 2 index files (orf_coding.fasta.*.bt2)

[zizigolu]

Sorry, but after indexing there are coding.1.bt2, coding.2.bt2, coding.3.bt2, coding.4.bt2, coding.rev.1.bt2 and coding.rev.2.bt2.

and orf_coding.fasta in bowtie2 file. Then what I should provide?

[Devon Ryan]

Then you need to write coding rather than orf_coding.fasta.

[zizigolu]

Thanks a lot Devon, you saved me I was getting mad since yesterday.