Evaluation of a Yeast Two-Hybrid Library by High-Throughput Sequencing

Yeast two-hybridization (Y2H) is
a classical method to study protein–protein
interactions in organisms, and the top limitation of Y2H is the high
ratio of false negatives and false positives. The most efficient way
to reduce this error is to improve the quality of the library. The
traditional library quality evaluation method can only inform us of
the capacity of the library and a very limited number of library insert
lengths. Therefore, we developed a new method to evaluate the library
by using only a 10 ng library, amplifying the inserted fragments through
15 cycles of PCR, and then carrying out high-throughput sequencing.
This method can eliminate the randomness and one-sidedness of the
traditional method and can be used to obtain key indicators, such
as the number of inserted genes and gene abundance, to effectively
evaluate the quality of the library. In addition, the new library
quality assessment method can also reveal the gene sequences of species.
This method is expected to greatly accelerate PPI research on nonmodel
species.