Hi Dear, I want to run a de novo transcriptome project by Roche 454 with a species of fish cDNA library that its genome size is about 3*10^9, which kind of picotiter plate region is better for me and why? 1/2 or 1/4 or 1/8 ?
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If the genome is 3E9 bp (similar to Homo sapiens)), then the transcriptome will be at least 3E6 bp. I'm not sure even 1/2 plate will have enough yield to cover your transcriptome, as there will be > 1000x difference between some transcript yields. You may need to use Illumina or Proton instead. -
I have run a devo transcriptome assembly using 454 on a fish with a similar sized genome (slightly larger than human genome). I ran 4 individuals in a whole 454 FLX run.
i used 3' targeted cDNA (however I seem to remember reading that for some reason this isnt the ideal protocol when using 454...?).
If memory serves I got around 30,000 contigs when aligning to a closely related reference transcriptome, and a bunch more via de novo assembly. From the 30, 000 contigs there were ~50K SNPs/indels identified....around 9K of these suited my criteria for a high quality putative SNP.
In theory 3' target cDNAs should increase yield of homologous fragments between individuals, and a full 454 plate seemed to give us good representation of the transcriptome.Last edited by JackieBadger; 10-07-2012, 11:43 AM.Comment
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