Transcriptome assembly and estimating abundance with Trinity
Hello,
I just want to check my experiment design with you:
I was given RNA-SEQ samples from some genetically engineered bacteria. So those bacteria contain their basic genome and an insert (pET plasmid). Goal: quantify the expression of the plasmid.
What I've done:
Bowtie2: aligned the reads to the reference genome; extracted unaligned genes.
Trinity: created a transcriptome assembly using the unaligned reads (assuming they are the ones corresponding to the plasmid).
Trinity: Align reads [Bowtie2] and estimate abundance where I'm feeding it the transcriptome assembly from step 2 and the RNA-SEQ data.
Does it make sense?
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Why not just including the plasmid sequence (which is known) to the reference genome?
I don't have the plasmid sequence. Basically, I'm trying to work with the unaligned reads.
Then ask the people who made the bacterial strain. It is always recommended to talk to collaborators first before asking strangers on the internet.
It's nothing to do with my collaborators. It's my first time using Trinity and I was curious how it works.
Plus, let's say I have the plasmid sequence. I still have to create an annotation file.