I want to express eukaryotic genes and yeasts seem a good bet to construct a library in. S. cerevisiae can't handle exons or introns, but I have not found definitive information about if Pichia pastoris does. Is there any other yeast that I should look into?
Edit: Thanks Kristof, I am hoping to express genes from other yeasts and possibly more diverse sets of fungi. I am hoping to avoid cDNA libraries because I am not sure that my targets are highly expressed. Thanks for the answer and the citation!
We had no problems in cloning of eukaryotic genes in P. pastoris, but the genes that we have cloned in P. pastoris had no introns
Dear,
Pichia pastoris is certainly an interesting tool for expression and allows cloning of libraries. Concerning your intron question, in the genome sequence of Pichia pastoris we have found a number of genes with one or a few introns (http://bioinformatics.psb.ugent.be/genomes/view/Pichia-pastoris) and the introns contain the common intron elements (donor, acceptor and branch point). Further, the introduction of a yeast intron in the sequence of hAAT1 increased the yield of hAAT1 significantly (http://www.ncbi.nlm.nih.gov/pubmed/16900336). So yes, Pichia pastoris can handle introns.
The question is what kind of eukaryotic genes do you mean, are these genes with a low number of introns or a high number of introns? In Pichia, the number of introns found in a gene is relatively low when compared to e.g. humans. Are these introns similar in size and structure of the Pichia introns?
Is it really needed to have the introns in the sequence? introns will increase the size of the genes which will make it harder to clone. If you could start from a cDNA library it will make your life easier.
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