[GenoMax]

Phred+64 refers to old format illumina (v.1.3+) quality calls. If you have recent data then it is going to be in Phred+33 (Sanger) format. See: http://en.wikipedia.org/wiki/FASTQ_format

[Gonza]

thanks. So if you have RNAseq data from an Hiseq 2500 then you should specify Phred+33 in tophat?

[GenoMax]

That is the default.

[Gonza]

great, thanks again!
g

[Nanu]

I exceuted the tophat to align against the reference genome. I found the following files:
1. accepted_hits.bam
2. unmapped.bam
3.junction.bed
4. align.txt

here accepted_hits.bam, size is 1 KB while unmapped.bam has 200MB. Align.txt shows :
Reads:
Input : 528840
Mapped : 0 ( 0.0% of input)
0.0% overall read mapping rate.
Please help me to define the parameters to align and mapped against the heterologous genomes.

[Nanu]

I download the data from Ensembl. Is it any problem in it.
I exceuted the tophat to align against the reference genome. I found the following files:
1. accepted_hits.bam
2. unmapped.bam
3.junction.bed
4. align.txt

here accepted_hits.bam, size is 1 KB while unmapped.bam has 200MB. Align.txt shows :
Reads:
Input : 528840
Mapped : 0 ( 0.0% of input)
0.0% overall read mapping rate.
Please help me to define the parameters to align and mapped against the heterologous genomes.