Yeast Hybridization Technology Service

After years of development, Tek Biotech has accumulated rich experience in immunology research and detection, and has rich experience in yeast library construction. Based on this, it provides high-quality yeast hybridization technology services to global customers, including yeast cDNA library construction technology services, yeast single hybridization technology services and yeast two-hybridization technology services.

█ Types of Yeast Hybridization Technology Services

TekBiotech has established a yeast hybridization verification system with the help of a mature yeast surface display technology platform. The principle is basically similar to that of antibody yeast surface display technology. Yeast hybridization technology can be divided into yeast single hybridization, yeast two-hybridization and yeast three-hybridization according to the type of detection. TekBiotech provides customers with yeast single hybridization and yeast two-hybridization technology services.

-- Yeast cDNA Library Construction Service

There are two conventional methods for constructing cDNA libraries: SMART technology and Gateway technology. Tianjin TekBiotech is good at using the SMART method combined with homologous recombination (homologous method of Gateway method) to construct cDNA display libraries. As shown in Figure 1, the SMART method synthesizes cDNA library:

Figure 1 Schematic diagram of SMART method for synthesizing cDNA library

For more information on yeast cDNA library construction technology, please refer to yeast cDNA expression library construction service

-- Yeast One-hybrid Technology Service (Y1H)

The yeast one-hybrid system belongs to the intranuclear interaction verification system, which is a gene reporting system built with the help of the biphasic functional characteristics of some cell transcription activators. In specific experimental operations, the transcription activator GAL4 is often used as a bridge to connect the upstream and downstream signal transduction. The basic principle is that the transcription factor GAL4 can regulate the expression of yeast galactosidase gene. When yeast expresses galactosidase, galactosidase catalyzes X-gal to produce blue colonies on the plate containing X-gal.

As shown in Figure 2, the transcription factor GAL4 contains two domains that can independently play a functional role: the transcription activation domain (AD) and the DNA binding domain (BD). The library containing various cDNA sequences was cloned into an expression vector with GAL4-AD to form vector 1; the cis-acting element was cloned into a vector with GAL4-BD to form vector 2. Then vector 1 and vector 2 are transformed into the same yeast cell for co-expression: if the fusion protein expressed by cDNA-GAL4-AD in vector 1 can bind to the cis-acting element in vector 2, the AD of GAL4 in vector 1 and the BD in vector 2 will combine to form a complete transcription factor GAL4, and GAL4 will bind to the upstream of the galactosidase gene, prompting the expression of the galactosidase gene to produce enzyme catalysis (i.e., reporter gene).

Figure 2 Principle of yeast one-hybrid system

For more information about yeast one-hybrid technology, please refer to yeast one-hybrid technology service

-- Yeast two-hybrid system (Y2H)

Similar to yeast one-hybrid technology, yeast two-hybrid technology uses the transcription activator to have two different domains: DNA binding domain (DNA-BD) and transcription activation domain (activation domain). The above two structures can function independently without affecting each other, but only when they are close enough in space can they exercise the activity of complete transcription activators, so that the downstream genes regulated by them can be transcribed. In the yeast two-hybrid system, the DNA-BD domain and the transcription factor AD domain are fused with X (bait protein) and Y (prey protein) to form fusion proteins. The DNA-BD domain-X fusion protein can recognize and bind to the upstream activator sequence (UAS: Upstream Activation Sequence) of the reporter gene. If the DNA-BD domain-X fusion protein interacts with the prey protein Y, this will make the DNA-AD domain and the BD domain approach each other and recombine to form a complete protein with the function of activating the reporter gene, and finally let the reporter gene express, as shown in Figure 3:

Figure 3 Principle of yeast two-hybrid system

Tianjin TekBiotech provides customers with two forms of yeast two-hybrid technology services: yeast two-hybrid technology (nuclear system) service and yeast two-hybrid technology (membrane system) service to meet the different project needs of customers.

（1）Yeast two-hybrid technology (nuclear system) service

Figure 4 Schematic diagram of yeast two-hybrid technology (nuclear system)

As shown in Figure 4, yeast two-hybrid technology (nuclear system) is similar to yeast one-hybrid technology, and uses the transcription activator GAL4 for library construction and screening. The library containing various cDNA sequences is cloned into an expression vector with GAL4-AD to form vector 1; the cis-acting element is cloned into a vector with GAL4-BD to form vector 2. Then vector 1 and vector 2 are transformed into the same yeast cell for co-expression: if the fusion protein expressed by cDNA-GAL4-AD in vector 1 can bind to the cis-acting element in vector 2, the AD of GAL4 in vector 1 and the BD in vector 2 will combine to form a complete transcription factor GAL4, and GAL4 will bind to the upstream of the galactosidase gene, prompting the expression of the galactosidase gene to produce enzyme activity catalysis (i.e., reporter gene).

For more information about yeast two-hybrid technology (nuclear system), please refer to yeast two-hybrid technology (nuclear system) service.

（2）Yeast Two-hybrid Technology (Membrane System) Service

Yeast two-hybrid technology (membrane system) is used to verify the interaction between membrane proteins and cytoplasmic proteins. The membrane protein interaction yeast two-hybrid system is based on the split-ubiquitin mediated separation-merge-cleavage system. The ubiquitin protein is divided into two domains: the N-terminal domain (called: Nub domain) and the C-terminal domain (called: Cub domain). These two domains can form a complete ubiquitin protein when they are close to each other inside the same cell. The ubiquitin protein can be recognized by ubiquitin-specific proteases (UBP, ubiquitin binding proteins; this enzyme recognizes the complete ubiquitin protein). The basic principle is shown in Figure 5:

Figure 5 Schematic diagram of membrane protein interaction yeast two-hybrid system

For more information about yeast two-hybrid technology (membrane system), please refer to yeast two-hybrid technology (membrane system) service

█ Advantages of Yeast Surface Display Technology Platform Service

Direct service from technical  experts to meet different  testing needs	Reliable results, cost-effective	High testing service efficiency,  real-time process monitoring	High sensitivity of testing  instruments, low sample  consumption	Short experimental cycle,  saving waiting time	Provide one-stop service: from  library construction to positive  clone screening, improve  efficiency, avoid errors,  and save costs