Chapter title |
Using Two-Hybrid Interactions to Identify Separation-of-Function Mutations.
|
---|---|
Chapter number | 9 |
Book title |
Yeast Genetics
|
Published in |
Methods in molecular biology, January 2014
|
DOI | 10.1007/978-1-4939-1363-3_9 |
Pubmed ID | |
Book ISBNs |
978-1-4939-1362-6, 978-1-4939-1363-3
|
Authors |
Brian Haarer, David C Amberg, David C. Amberg, Haarer, Brian, Amberg, David C. |
Abstract |
Protein functions within cells frequently require they interact physically with a number of partner proteins to coordinate the appropriate biochemical processes. Mutational analysis has been quite useful for analyzing how the loss of a gene or protein impacts cell function or more specifically particular pathways. However, the genetics approach to studying gene function can be limited by not having the right mutations; for example because the mutant allele ablates all function, as is the case for deletion (null) alleles and most temperature-sensitive alleles. To dissect the relative contributions of a protein's interactions, the researcher needs mutations that specifically affect one but not all of the protein's interactions. In genetics parlance such mutations are called separation-of-function mutations. The yeast two-hybrid system has been exploited for two decades to identify protein-binding partners. Here we describe a fairly simple protocol, within reach of laboratories with molecular biology experience, for using the two-hybrid system to identify separation-of-function mutations. |
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