When you assembly a transcriptome with Trinity, for example, only one final fasta is created with the transcriptome. The de novo transcriptome assembly does not assemble transcripts for the separate alleles, and usually there is only one transcript generated and it is mapped to both alleles. Is there any software that allows assembly de novo and with reference genome transcrips for separete alleles?
If you want haplotype aware assembly you need long reads. With short NGS sequences you don't know when the haplotype ends. To work around this you can map back your NGS reads to the assembly and see which allele is more common, ie. that there is some allele specific expression change.
But if there are multiple alternative transcripts I doubt that short read sequencing will be helpful in determining if particular allele is a part of this or that alternative transcript.
Dear Raquel,
it sounds kind of contradictory. On the one hand, you do de novo transcriptome assembly. On the other hand, you'd like to map to reference genome transcripts. Typically, you would do the de novo assembly only because a reference genome or reference transcripts are not available. In case, you have references beforehand, read mapping to genome or transcriptome would just solve it.
In case of lacking references, I recommend the basic strategy:
- de novo assembly, as you did
- identification of potential allele variation (SNVs). The assembly graph generated by Trinity can be used to do this. Alternatively, transcript mapping and subsequent variation analysis would reveal the variation.
- final mapping to separate the alleles and quantify the allele variants.
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