Blue Light Controllable Yeast System

Chemical regulatory systems are easy to use and can produce high levels of protein expression, but they usually provide imprecise control of protein levels. Light is dose-dependent, low application cost, easy to deliver, and will not interfere with cellular pathways, so it has significant advantages over chemical systems.

Creative BioMart can now insert photoreceptors and their interacting proteins into the transcriptional activation elements of the target gene, so as to control the GAL1 promoter and the transcription of GOI using the blue light / cryptochrome dependent system

Construction of the Blue Light Controllable Yeast System

The switch is based on a light activated yeast two hybrid system, in which the blue light photoreceptor is connected to the DNA binding domain (DBD), and the photoreceptor interaction protein is fused to the trans activation domain (AD).

The system is based on blue light stimulated photoreceptor cryptochrome 2 (CRY2) and its interaction partner CIB1 Blue light can bind CRY2–CIB1 to aggregate the split Gal4 transcription factors that control DNA transcription. The dark reversion of CRY2 separates the interaction with CIB1 and stops Gal4 dependent transcription.

Creative BioMart has been engaged in microbial gene editing for many years and has a perfect Saccharomyces cerevisiae gene editing technology platform. Our gene editing technology platform has experienced technicians, efficient experimental process and perfect experimental equipment. Our scientists can customize one-to-one solutions according to the specific needs of customers, and provide editing services for a variety of microorganisms to meet the needs of customers.

Recombinant Protein Expression System

Blue light controlled protein expression has the following advantages

The transmission of light is immediate and does not require time to absorb chemicals into cells
Light does not need to operate the medium, and can be easily automated
Light is minimally invasive
Light is not affected by compound absorption or drug pump problems
The light induction system allows precise dose-dependent control of protein levels because light can be transmitted within a specified time and removed immediately

Step	Description	Deliverable	Time
Subcloning of yeast expression vector	Codon optimization, gene synthesis (or customer provided plasmid template: target gene amplification) The target gene was subcloned into a suitable expression plasmid Splice sequence validation: sequencing	Correctly constructed recombinant expression plasmid, DH5 containing recombinant plasmid α Strain and sequencing original report	1-2 weeks
Yeast expressing strain transformation	Amplify and culture 5-10 positive clones and induce the expression of recombinant protein	SDS-PAGE delivery: report of positive clone strains and expression screening results.	2 weeks
Screening of yeast expressing strains	Detection of recombinant protein expression (applicable to multi copy number and high expression strains)	Report of positive clone strains and expression screening results.	2 weeks
Optimization of yeast expression strains	For the selected monoclonal strains, the culture and induction conditions (medium, cell density, methanol concentration, induction time, etc.) were optimized to improve the expression of the target protein		2-3 weeks
Small scale protein expression and purification	1L fermentation broth was purified by affinity chromatography, ion exchange, hydrophobicity and gel filtration.	SDS-PAGE delivery: recombinant protein and purification report. The final protein containing purified tag or excised purified tag can be provided according to customer requirements, with purity > 85%.	2-3 weeks
Large scale protein expression and purification	10L, 30L, 80L, 130L, 500L fermentation scale.

Creative BioMart is always devote us to provide high-quality and satisfactory service to our customers, if you are interested in our services or have some question, please feel free to contact us or make online inquiry.