Creative Biolabs provides yeast two-hybrid services for the discovery or validation of new interactions. For soluble or membrane molecules, we have optimized systems for Y2H library construction and screening.
All processes in living cells virtually depend on protein-protein interactions (PPIs). Thus, it is essential for molecular biology and disease research to understand PPIs networks. Yeast two-hybrid (Y2H) is an extremely powerful technique for studying protein-protein interactions and especially, can be used to search for a novel interacting partner by screening a single protein or domain against a library of other proteins. It is this latter characteristic that to find the interacting protein without any prior identity of such proteins is the strongest application of Y2H assay.
Fig.1 A simple flow-chart for a random yeast two-hybrid (Y2H) library screening.
Y2H as a cogent genetic system is one of the most standardized approaches currently for mapping PPIs both at a small scale and in a high throughput manner. In Y2H screening, PPIs are detected by the activation of reporter genes which respond to a reconstituted transcription factor (TF). Y2H is commonly applied to test some “prey” proteins for interactions with a single “bait” or target protein or pool of proteins. The merit of this method is the direct identification of interacting protein pairs without downstream experiments since the prey protein has been known and do not need further confirmation.
Rationale of Y2H
Y2H was originally developed by Stanley Fields and Ok-Kyu Song in 1989. The rationale of this system is dependent on the physical separation of the DNA-binding domain (DBD) from the transcriptional activation domain (AD) of several transcription factors. Y2H is enabled by the fact that the domain of a fully functional protein can be separated and then recombined to constitute full functional protein.
Fig.2 The yeast two-hybrid system is a genetic tool used to detect interactions between two proteins.
Specifically, a TF can be split into a DBD and an AD. In a typical Y2H, a protein “X” of interest is fused to the DBD of the yeast transcription factor Gal4. This fusion of DBD-X is generally referred to as a bait protein. The DBD is able to bind to the upstream activating sequence of a promoter but only activates the transcription when an AD is present. Meanwhile, random protein fragments or open reading frames (ORFs) are fused to the AD of Gal4, which fusions are called preys. The AD has the ability to activate transcription but cannot bind to the promoter of a reporter gene without a DBD. Therefore, only if the bait and prey are interacted and co-expressed, a fully functional TF will be reconstituted and subsequently, the reporter gene transcribed.
Advantages and Key Features of Y2H Service
Creative Biolabs provides the services on a strict fee-for-service basis. We are professional to use Gal4 and LexA based yeast two-hybrid (Y2H) systems.
Creative Biolabs optimized Y2H protocols. The efficiency and quality of our Y2H system have been satisfying clients with highly reproducible and reliable results. We are also skilled at other types of two-hybrid programs, such as mammalian two-hybrid and bacterial two-hybrid services, to suit worldwide scientists’ need in a wide range of biological fields. Please contact us for more details.