pGBK/pGAD false positives in two-hybrid

Ricky Boernke ricky_boernke at gmx.net
Thu Sep 2 02:23:59 EST 2004


Dear Yeasters,

We've been using the yeast two-hybrid system based on pGBT-9 and
pAD-GAL4 (from Stratagene) in past and have been pretty successful
with this combination. Lately, we switched to Clontech's Matchmaker 3
system since their vector system appeared to offer some advantages
over the old system. However, it seems that screening using baits in
pGBK-T7 give rise to loads of false positives which we never picked up
before from our library and this is independent of the bait protein we
use. In turn, screening a library constructed into pGAD-T7 using the
SMART library construction kit with bait proteins in pGBT-9 also picks
up loads of the same fals positives. So my feeling is that something
in those MM3 vectors is responsible for that phenomenon.  Might be the
high expression level or the HA/myc-tag. Have you observed this
before?

Best wishes
Ricky

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