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Which library prep kit for targeted RNA-seq of an overexpression library?
I am going to transform a library of overexpression constructs into my cells, and then after applying a selection pressure I want to do targeted RNA-seq to determine the relative expression of each library gene in the pool of selected cells.
The vector backbone is the same for all constructs in the library, and therefore every transcript will have the same 5' UTR and 3' UTR, the latter of which contains a designed primer binding site for first strand cDNA synthesis. I only want to check the expression of the library genes; I do not want to sequence the transcriptome. The size of the genes in the library range from about 200-1200 bp. All of the library genes have very different sequences, so I only need to sequence about 75-150 bp in order to be able to map the reads to the reference sequences.
Which Illumina library prep kit should I use and why? Or should I instead use a custom protocol since I have a designed primer site for first strand cDNA synthesis?
Thanks for your help.
The vector backbone is the same for all constructs in the library, and therefore every transcript will have the same 5' UTR and 3' UTR, the latter of which contains a designed primer binding site for first strand cDNA synthesis. I only want to check the expression of the library genes; I do not want to sequence the transcriptome. The size of the genes in the library range from about 200-1200 bp. All of the library genes have very different sequences, so I only need to sequence about 75-150 bp in order to be able to map the reads to the reference sequences.
Which Illumina library prep kit should I use and why? Or should I instead use a custom protocol since I have a designed primer site for first strand cDNA synthesis?
Thanks for your help.