Tophat2 alignment for csfasta file

Question

[nikkihathi]

Hello!

I have the same problem for tophat to recognise the index file for the genome. I don't understand the problem.

I am using the following command

  tophat -p 6 --color --bowtie1 --quals --library-type fr-secondstrand -G /Genomes/mouce_database_10/mm10_c data1-RNA.csfasta data1-RNA.QV.qual

and this give the error!Traceback (most recent call last):

 File "/usr/local/tophat/tophat", line 4095, in <module>
 sys.exit(main())
 File "/usr/local/tophat/tophat", line 3869, in main
 left_quals_list = args[2]
 IndexError: list index out of range

I check the location and number of files available in Genome folder. I found 6 .ebwt files in the Genome folder. To make genome index file, I used

 bowtie-build -C  mm10.fa mm10_c

Is there any command that could be useful to solve this problem? and what does this error message means? any help is appreciated.

Thanks

Answer 1

[EagleEye]

I had same problem, tophat2 only works with bowtie2 index,

1) the bowtie1 index is not supported by tophat2

2) you cannot build index for color space reads using bowtie2 because bowtie2 build does not support color space read anymore(-C option).

3) the solution is,

A) build index using Bowtie1

B) align with tophat1 version

C) or try alternate aligners like Shrimp, subread etc.,

It worked for me.

[nikkihathi]

Thanks for your reply and suggestion for the steps to follow for the alignment. I am looking for a pipeline to predict fusion genes in color space reads as in Tophat there is an option for fusion search.

Also, I would like to ask if there are any quality control measures that I should be looking into for color space reads? If you could suggest paper or blogs that have discussed this issue

[EagleEye]

1. Sorry I am not familiar with fusion gene analysis.

2. I can hardly think of any blogs about color space reads. But you can use adaptor removal tool Cutadapt (supports color space reads) as initial quality control measure.

[nikkihathi]

Thanks for your reply.