Multiple index files in Bowtie?

[spb2003]

Hello Everyone,

I am working on a 32 bit RH Linux machine and setting up a RNASeq analysis pipeline. As part of ERANGE, I am building an index of the expanded genome but on including .fa.masked files the total number of characters balloons to more than 2^32. I went ahead and built two index files by splitting all the files into two sets.
I tried an small example of aligning a 36bp read against both the refseq and ensembl index. No alignments are found if I use two index together in the command line but they are found when searched against one index at a time. Is this the only option when there are multiple index files or am I missing something more obvious?

Thanks for your help.

[GeeSharpMinor]

I run an RNAseq pipeline where we sequentially align against two different indexes, maybe I can help. can you copy/paste the commands you use, also listing any options?

[spb2003]

for eg. I want to align CCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA against two index files hg18_block1 and hg18_block2. Since I am still building the index, I will give an example using different index

$bowtie -t human_ensembl_cdna -c CCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA
Time loading forward index: 00:00:01
Time loading mirror index: 00:00:02
Seeded quality full-index search: 00:00:00
0 + ENST00000322310 755 CCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 0 7:T>C,18:C>N
Reported 1 alignments to 1 output stream(s)
Time searching: 00:00:03
Overall time: 00:00:03

$bowtie -t refseq_human -c CCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA
Time loading forward index: 00:00:02
Time loading mirror index: 00:00:01
Seeded quality full-index search: 00:00:00
0 + REFSEQ:NM_003731 800 CCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 0 7:T>C,18:C>N
Reported 1 alignments to 1 output stream(s)
Time searching: 00:00:03
Overall time: 00:00:03

The following does not work:
$bowtie -t human_ensembl_cdna -t refseq_human -c CCCTCACCCTGTCCCTCCTNAGCCAGCAGAGGCCCA
Time loading forward index: 00:00:02
Time loading mirror index: 00:00:01
Seeded quality full-index search: 00:00:00
No results
Time searching: 00:00:03
Overall time: 00:00:03

Is a sequential search the only option or is there a way to do alignment against multiple index files simultaneously.

Thanks.

[GeeSharpMinor]

unfortunately, sequential is the only way to do it as far as I know.