Yeast One-hybrid Assay (Y1H)
Technical principle:
Transcription factors typically consist of a DNA-binding domain (BD) that specifically binds to DNA and one or more activation domains (AD) that interact with other regulatory proteins. In the yeast one-hybrid system, a typical transcription factor is the yeast GAL4 protein. The BD of GAL4 is near the carboxyl terminus and can activate the upstream activation site (UAS) of yeast galactosidase, while the AD can interact with RNA polymerase or transcription factors (TF), enhancing the activity of RNA polymerase. BD and AD can function entirely independently, allowing the BD of GAL4 to be replaced by genes encoding library proteins. As long as the expressed protein can interact with the target gene, the AD will similarly activate RNA polymerase, initiating the transcription of downstream reporter genes.
Delivery time
|
Experimental steps |
Delivery Time(days) |
Delivery Materials |
|
A. Synthesis and amplification of bait plasmid |
15-20 |
|
|
B. Linearization of bait plasmid |
10-15 |
|
|
C. Integrate the linearized bait plasmid into Y1H |
||
|
D. Identification, sequencing, and alignment analysis |
||
|
||
|
F. Preliminary screening |
15-20 |
|
|
G. Repeat screening |
||
|
H. Positive clone sequencing |
||
|
I. One-to-one verification |
15-20 |
|
|
||
|
Our Service Advantages:
Library Selection:
- Customized yeast library with four uses (single hybrid, double hybrid, triple hybrid, overexpression vector library): Uses optimized Clontech's pGADT7-Rec2, GATEWAY secondary recombination method, and 'one-step recombination' library;
- Provides commercial yeast cDNA libraries.
Screening Service:
- The yeast nuclear system single-hybrid system can study interactions between DNA and proteins, with optimized screening steps and a mature experimental workflow, providing reliable experimental data and original images.
- High-throughput screening with Y1H-seq can be conducted after screening.
- Two bait vector screening options: pHIS2 or pAbai vectors.
