de novo assmebly of RNA data using Velvet

[santhosh003]

Hi,
I have Illumina RNA single-end read data with 4 million sequences of read size 33 bases each. I used Velvet to perform de novo assembly on the RNA data. Velvet has generated contigs for the same. Since I come from computing background and has very little biology knowledge, I would appreciate if somebody can help me with the next steps. Does the output of velvet is final? What should be my confidence level looking at the contigs? What should do to verify/validate the output contigs? Should I use BLAST to find genes? Or can anybody suggest with a general pipleline that is followed to perform analysis post de novo assembly. Any help in this matter is greatly appreciated. Thank you.

[Xi Wang]

You may use Oases, which is more applicable for RNA-seq data than Velvet.
The link:
http://www.ebi.ac.uk/~zerbino/oases/

[santhosh003]

Hi XI Wang,
Thanks for the reply. I have used Oases, but i want to know what exactly is the output file among the files that it has created? Is there any viewer like "MAQ View" to view the output in a better way? Thank you.

[Xi Wang]

Hi XI Wang,
Thanks for the reply. I have used Oases, but i want to know what exactly is the output file among the files that it has created? Is there any viewer like "MAQ View" to view the output in a better way? Thank you.

MAQ view is viewer for sequence alignment, right? So to do this, you need to introduce a additional step, that is to align the assembled transcripts to the reference genome. The output files of Oases contain a fasta file for the transcripts' sequences. This alignment is somehow similar to the way aligning EST sequences to the genome, so you can use the tool BLAT, which allows you to align with several mismatches and gaps.

BTW, my surname is Wang, and the given name is Xi.