I have four transcriptome assembly of the same species but with different treatments . I want to annotate each of transcriptome assembly to closely related reference cDNA species (from ensembl). Then I want to compare each of the annotated transcriptome assembly and to see how many genes appear.
I want to know which genes present in all treatments and what are all the genes that are absent in different treatments. I have tried tblastx with ensembl (cDNA) sequence against transcriptome assemblies.Trying to find out genes which having highest blast coverage.I would like to get idea to better way to compare to get list of genes. Any journal paper info regarding this will be helpful.
Does your species have a reference genome? Is so, you can get your assembled transcripts from each treatment into GTF format and a reference GTF transcriptome file from Ensembl such as those available on their FTP site, you should be able to use Cuffcompare for these kinds of comparisons. Pay particular attention to the '-r' option. It will assign a class code that describes how each assembled transcript compares to your reference transcripts as folllows:
Class Codes
Priority Code Description
1 = Complete match of intron chain
2 c Contained
3 j Potentially novel isoform (fragment): at least one splice junction is shared with a reference transcript
4 e Single exon transfrag overlapping a reference exon and at least 10 bp of a reference intron, indicating a possible pre-mRNA fragment.
5 i A transfrag falling entirely within a reference intron
6 o Generic exonic overlap with a reference transcript
7 p Possible polymerase run-on fragment (within 2Kbases of a reference transcript)
8 r Repeat. Currently determined by looking at the soft-masked reference sequence and applied to transcripts where at least 50% of the bases are lower case
9 u Unknown, intergenic transcript
Hi Malachig.......i was actually trying to do the same. The problem i had is i was not been able to convert my blastx results to GTF. i tried doing blastx to gff but from gff to gtf i am having hard time. Could you please point to me scripts/tools that can convert my blastx results to GTF so that i can use it with cuffcompare. Also i want to ask... when you convert your assembled transcripts into GTF do you blast them against your reference genome or transcriptome?
Thanks for your reply. My species doesn't have reference genome. But I have Ensembl cDNA sequence. Is any other way to compare.
Hi Malachig.......i was actually trying to do the same. The problem i had is i was not been able to convert my blastx results to GTF. i tried doing blastx to gff but from gff to gtf i am having hard time. Could you please point to me scripts/tools that can convert my blastx results to GTF so that i can use it with cuffcompare. Also i want to ask... when you convert your assembled transcripts into GTF do you blast them against your reference genome or transcriptome?