DUALhybrid Protein Interaction Kit

Detect Protein Interactions in vivo

Classic but with unique improvements

The DUALhybrid kit for cytosolic and nuclear proteins is based on the classical yeast two-hybrid system and includes a range of improvements to facilitate yeast two-hybrid screening. The DUALhybrid technology enables you to screen a protein of interest for interactors expressed from a cDNA library of your choice. Two different bait vectors give you the option to express your protein of interest either as N- or C-terminal fusion to the LexA domain. The choice of two orientations enhances your chances of finding all interactors for your protein and reduces the chances of self-activation. The triple reporter strain NMY51 allows high stringency selection and greatly lowers the rate of false positives in a screen.

The technology

A bait is constructed by fusing a protein of interest to the LexA domain. The LexA domain localizes the bait to a particular DNA sequence upstream of a reporter gene.

A prey is constructed by fusing a second protein of interest to the activation domain of a transcription factor.

An interaction between the two proteins recruits the prey to the start of the reporter gene and the hybrid transcription factor assembled in this way from bait and prey (hence “two-hybrid”) activates the reporter gene. A protein interaction is thus measured by reporter gene activity. Commonly used reporter genes are either auxotrophic markers which allow selection on a minimal medium or color reporters like lacZ.

Screening of cDNA libraries to identify novel interactions

The defined prey can be replaced by a collection of different preys (i.e. different proteins fused to the activation domain) expressed from a cDNA library. Each yeast cell expresses the same bait and a particular prey from the cDNA library. If bait and prey interact, the yeast cell multiplies and gives rise to a visible colony on minimal medium. All yeast cells which do not express an interacting bait/prey pair fail to grow and are discarded. Finally, yeast colonies are picked and the prey plasmid is isolated and sequenced, yielding the information on the protein which interacts with the bait. The final result of a yeast two-hybrid screen is a list of proteins which interact with your protein of interest (the bait). It is important to realize that standard cDNA libraries can over-represent abundant mRNAs, and this can lead to skewing screens toward those proteins which are expressed at higher levels. That is why normalized libraries are offered, so that only the most relevant interactions are selected. The normalized cDNA libraries are constructed utilizing reassociation kinetics to target and reduce over-abundant cDNAs. The net result can greatly decrease the rate of non-specific interactors and increase chances of identifying rare interactors.

The odds are stacked in your favor

What separates Dualsystems protein-interaction screening technology from home brew screens or other company kits? Each step has been optimized such that relevant interactors are identified in 98% of all screens. Traditional pitfalls have been addressed by technological advances like normalized cDNA libraries, C-terminus vs. N-terminus baits, and location-specific interactions. Dualsystems products have been used in hundreds of screens annually for the past ten years, and their vast knowledge has been applied to making state-of-the-art systems that promise the best for in vivo analysis of protein interactions. More than sixty peer-reviewed papers, over the past 8 years, refer to their products.

DUALhybrid Kit

ITEM #	SIZE	DESCRIPTION	PRICE	BUY
P01004	1 Kit	DUALhybrid kit	$1010

See our DUALhybrid normalized cDNA library and non-normalized cDNA collections.

Vector Maps and Sequences

ITEM #	VECTOR	DESCRIPTION	PRICE	MAP	SEQUENCE
P03105	pLexA-N	Bait vector for expression of cDNAs fused to the C-terminus of LexA.	$430
P03106	pLexA-C	Inverse bait vector for expression of cDNAs fused to the N-terminus of LexA.	$430
P03103	pGAD-HA	Library vector based on the popular pGAD series.	$430
P03104	pY3H	Bridging vector for the expression of a third protein partner in the yeast two-hybrid system.	$430
P03107	pLexA-p53 pLexA-laminC pACT-largeT	DUALhybrid control plasmid set (3 controls)	$430