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Customized Enzyme
The CRISPR-Cas system, known for its unique ability to recognize target nucleic acids and its efficient cutting capability, has been widely applied in gene editing and other fields. Cas nucleases are the key components of the CRISPR-Cas system, functioning as RNA-guided endonucleases capable of cleaving DNA and RNA molecules. Among the various types of Cas enzymes, Cas9, Cas12a, and Cas13a have been extensively utilized.
Service Details
| Applications | 1.Gene Editing 2.CRISPR-Based Detection |
|---|---|
| Timeline/Pricing |
 Consult online for details |
EDITGENE offers customized enzyme development services for gene editing, backed by extensive experience in Cas enzyme development and rigorous production processes. Utilizing an exclusive protein expression and purification platform, our service involves software-assisted design and optimization of gene editing enzymes, efficiently screening candidate enzymes. With our proprietary Fish/Bait one-step purification technology, we significantly simplify the protein purification process, reducing potential protein loss. This approach ensures precise design and high-purity gene editing enzymes for our clients through efficient screening.
Service Advantages
Extensive CRISPR Detection Experience
Custom enzyme design for CRISPR detection based on client requirements
Flexible Delivery Options
Various types and specifications available
Years of Gene Editing Experience
Custom enzyme design based on client needs
Technical Roadmap
Developed Product Categories/Types
| Spcas9 | Lbcas12a | Aapcas12b | Lbucas13a | Lwacas13b | |
|---|---|---|---|---|---|
| PAM/PFS | 5’-NGG-3’ | 5’-TTTV-3’ | 5’-TTN-3’ | None | None |
| None | dsDNA | dsDNA or ssDNA | dsDNA or ssDNA | ssRNA | ssRNA |
| Cis-cleavage | dsDNA | dsDNA or ssDNA | dsDNA or ssDNA | ssRNA | ssRNA |
| Trans-cleavage | None | ssDNA | ssDNA | ssRNA | ssRNA |
SpCas9 is an sgRNA-guided endonuclease that specifically binds and cleaves target dsDNA at the presence of a PAM sequence (5'-NGG-3'). Guided by sgRNA, SpCas9 cleaves the target DNA at a site located three nucleotides upstream of the PAM sequence, generating blunt ends. Besides gene editing, SpCas9 can be used for in vitro target DNA cleavage and cloning of desired fragments.
LbCas12a is a crRNA-mediated endonuclease that specifically recognizes and cleaves dsDNA in the presence of a PAM sequence (5'-TTTV-3'). Additionally, LbCas12a can cleave ssDNA without a PAM sequence. Upon binding to complementary dsDNA or ssDNA, the LbCas12a-crRNA complex activates its non-specific trans cleavage of ssDNA. By designing ssDNA reporters labeled with fluorescent tags or other molecules, LbCas12a can be used for DNA template detection and signal amplification, with results observable via fluorescence or test strips.
AapCas12b is an sgRNA-guided endonuclease that cleaves dsDNA at PAM sequences (5'-TTN-3'). AapCas12b also exhibits non-specific ssDNA trans cleavage after binding to complementary dsDNA or ssDNA. With an optimal reaction temperature of 60°C, AapCas12b is well-suited for use in conjunction with loop-mediated isothermal amplification (LAMP).
LbuCas13a is a crRNA-mediated endonuclease that is PAM-independent. Upon recognition and cleavage of target RNA, LbuCas13a's trans cleavage activity is activated, allowing non-specific cleavage of ssRNA. Detection and signal amplification can be achieved using RNA probes labeled with fluorescent tags or other markers, with results observable via fluorescence or test strips.
LwaCas13a, like LbuCas13a, is a crRNA-mediated endonuclease that is PAM-independent. When crRNA recognizes and cleaves target RNA, LwaCas13a's trans cleavage activity is activated, enabling non-specific cleavage of ssRNA in the system. CRISPR/Cas13a-mediated detection and signal amplification can be performed using RNA probes labeled with fluorescent or other markers, with results viewable via fluorescence or test strips.
Advantage and Characteristic
Optimazied Strategy
We have create a unique sgRNA Design Logic
Optimazied Strategy
We have create a unique sgRNA Design Logic
Optimazied Strategy
We have create a unique sgRNA Design Logic
Optimazied Strategy
We have create a unique sgRNA Design Logic
FAQ
What are the differences between Cas9, Cas12, and Cas13?
The main differences among Cas9, Cas12, and Cas13 lie in their action mechanisms:
· Cas12 is activated to cleave ssDNA trans-cleaving after binding with guide RNA and target DNA.
· Cas13 is activated to cleave ssRNA trans-cleaving after binding with guide RNA and target RNA.
· Cas9 has not been reported to exhibit trans-cleaving activity.
· Cas12 is activated to cleave ssDNA trans-cleaving after binding with guide RNA and target DNA.
· Cas13 is activated to cleave ssRNA trans-cleaving after binding with guide RNA and target RNA.
· Cas9 has not been reported to exhibit trans-cleaving activity.
How long can CRISPR-related reagents and Cas proteins be stored?
CRISPR detection reagents:
1.The RPA isothermal amplification kit can be stored at -20°C for long-term storage.
2.Target plasmids can be stored at -20°C for long-term use.
3.Cas proteins are sensitive to repeated freeze-thaw cycles; it is recommended to aliquot into multiple tubes and store at -80°C, retrieving them as needed for experiments. For short-term use, they can be stored at -20°C.
4.crRNA is prone to degradation and should be stored at -80°C if not used in the short term.
5. Probes, being double-stranded DNA, are relatively stable and can be stored at -20°C.
1.The RPA isothermal amplification kit can be stored at -20°C for long-term storage.
2.Target plasmids can be stored at -20°C for long-term use.
3.Cas proteins are sensitive to repeated freeze-thaw cycles; it is recommended to aliquot into multiple tubes and store at -80°C, retrieving them as needed for experiments. For short-term use, they can be stored at -20°C.
4.crRNA is prone to degradation and should be stored at -80°C if not used in the short term.
5. Probes, being double-stranded DNA, are relatively stable and can be stored at -20°C.
Can both dsDNA and ssDNA targets activate the trans-cleaving activity of Cas12a? Which has higher efficiency?
Both double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA) targets can activate the trans-cleaving activity (also known as collateral cleavage) of Cas12a, similar to Cas12b. However, the efficiency differs: ssDNA targets activate Cas12b trans-cleaving activity more efficiently than dsDNA targets, while dsDNA targets activate Cas12a trans-cleaving activity more efficiently than ssDNA targets.
