Hello Everyone,

I am planing to conduct a Yeast One-Hybrid screen using the Takara Matchmaker Y1H Gold screening kit.

Reading trough the manual I did not really find information on how this strategy would ensure, that the cDNA library clones are translated in frame with the fused Gal4 activation domain. There is only one section elaborating on this problem saying, that yeast can tolerate frame shifts and as I understand still expresses the right protein.

As I remember in the past there used to be vector systems, where you could insert your cDNA clones in different frames (e.g. +1, +2, +3) and therefore one of the three vectors resulted an in-frame protein fusion. (for clarity: in such case you had to prepare 3 libraries one with each vector)

Maybe there is a trick during the SMART cDNA synthesis. I mean that the adaptors which are fused to the cDNA library clones for recombination cloning are ensuring that every third of the cloned transcripts are cloned in a different frame.

I would appreciate if anyone can clarify this question for me.