454 - How to calculate frequency ?

[Giorgio C]

Hy,
i have assembled a transcriptome with newbler and than blast the results.
Do you know how to calculate or to find in any file the frequency of any isotig-isogroup so to see which are better represented ?
Thank you

[Jeremy]

The number of reads per contig is outputed. But, determining the distribution of reads is difficult for large isogroups. For isogroups with one isotig you can simply map (gsMapper works nicely) the raw reads against the isotigs.fna which will output a file with read counts per isotig. Isogroups with >1 isotig presents a problem though, do you map against all the contigs and combine the counts or map against one of the isotigs per isogroup (which will at least give read counts for a {probably} real mRNA)?

[Giorgio C]

Thank you for your answer. I would to estimate the most represented gene. So i though to blast all the sequences coming out from the 454 extracted from the .sff and converted in fasta (before assembly) against the "orf" database. So to identify the expression. What do you thin about?