- Identify critical amino acids required for your favorite interaction
- Correlate loss of interaction with loss of function in cells
- Full-length proteins or fragments, peptides
- Cytoplasmic or extracellular proteins
- Loops and tails of membrane proteins
How does it work?
Mutants are generated by PCR and the experimental conditions are adapted to each target gene to get on average a single mutation per clone.
The mutant library is then screened against the interacting partner by yeast two-hybrid using the LacZ reporter gene. White or light blue yeast clones in which no or a weak interaction occurs are selected.
192 clones are fully sequenced to identify their mutation. Their phenotypes are confirmed in a secondary, semi-quantitative LacZ assay.
You select up to 10 point mutants which are further characterized in a quantitative colorimetric interaction assay. You then receive DNA plasmids for these 10 mutants for your own susbsequent functional studies.