Hello everyone,
currently I´m trying to amplify a C. reinhardtii gene out of cDNA. It is not working so far. I tried different PCR methods and ordered also new primers with longer gene specific parts. Now I had the idea to amplify it from the genomic DNA (the gene has 9 exons). I could try to remove every single intron by GoldenGate cloning, but I thought there might be an easier way. Can I express this gene in A. thaliana protoplasts and extract the RNA of them to produce a lot of cDNA as template for my PCR? To do this I have to know, if Arabidopsis can recognize and splice the present introns in this gene correctly? I hope the PCR will work better with more specific cDNA as template.
Doubtful. Algae and land plants diverged millions of years ago.
Might want to look into when/where your gene of interest is expressed in C. renhardtii. If your transcript is very rare, that does make it more challenging to create your desired cDNA.
You could also consider having the gene synthesized if you're concerned about splicing issues.
- Badges
- Science method