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32122 SuCas12a2 Nuclease

  • 32122 SuCas12a2 Nuclease.png

32122 SuCas12a2 Nuclease

  • Specifications:
    • 100 pmoL
    • 1,000 pmoL

Article number:

32122


Price:Contact our distributors


隐藏域元素占位

Product overview

Description

SuCas12a2 nuclease is derived from the Sulfuricurvum sp. PC08-66 strain. Cas12a2 is a crRNA-guided RNA endonuclease, and its direct repeat sequence (DR) in crRNA is similar to that of Cas12a. Cas12a2 can specifically recognize and cleave the ssRNA target in the presence of a PFS sequence on the target ssRNA. In addition, compared to other Cas nucleases that, when specifically activated by the target, only non-specifically degrade ssDNA (e.g., Cas12a, Cas12b), or ssRNA (e.g., Cas13a), or both ssRNA and ssDNA (e.g., Cas12g), Cas12a2 has a more powerful trans-cleavage activity (i.e., collateral cleavage activity). This means that when Cas12a2 enzyme forms a ternary complex with crRNA and target RNA, it is activated to perform trans-cleavage activity against non-specific sequences of ssDNA, ssRNA, and dsDNA, cleaving any sequence of ssDNA, ssRNA, and dsDNA in the system.

 

 

Product Components

Components

32122-01 (100 pmol)

32122-03 (1,000 pmol)

SuCas12a2 Nickase (10 μM) a

100 pmol

1,000 pmol

10 × HOLMES Buffer 1

1 mL

1 mL

a.The concentration of SuCas12a2 Nickase is 10 μM (10 pmol/μL);

 

 

Storage Conditions

Store at -20℃; transport at ≤0℃.

 

 

Experimental Procedure

The nicking experiment: In order to verify that SpCas9 H840A Nickase can cleave the NT strand in the specific dsDNA target, we used supercoiled DNA (cccDNA) as the target. SpCas9 H840A Nickase, sgRNA, and target cccDNA were added to the cleavage reaction system, which was then incubated at 37°C for 30 min before being inactivated at 85°C for 5 min. The nicked open circular DNA (ocDNA) has a slower migration rate than the original cccDNA, so the sample bands can be detected by 1% agarose gel electrophoresis (150 V, 20 min). The experimental results showed that SpCas9 H840A Nickase generated a nicking site on target cccDNA under the guidance of sgRNA.

 

 

Experiment Results

 

 

Notes

 

1. Different sources of Cas12a2 enzymes may have slightly different requirements for the recognition of target PFS sites, with most Cas12a2 enzymes recognizing the 5′-GAAAG-3′ site.

2. Cis-cleavage of Cas12a2: Cas12a2, guided by crRNA, specifically cleaves Target RNA with PFS sites.

3. Trans-cleavage of Cas12a2: When Target RNA is present, Cas12a2, crRNA and Target RNA forms a ternary complex (Cas12a2/crRNA/Target RNA) and simultaneously activates the trans-cleavage activity of Cas12a2, allowing it to cleave any sequence in the reaction system, including ssDNA, ssRNA, and dsDNA.

4. To prevent RNase contamination, maintain a clean and tidy experimental area, and wear clean gloves and masks during operation. All consumables such as tips and centrifuge tubes used in the experiment should be RNase-free.

5. Cas12a2 enzymes are sensitive to heat and prone to inactivation. Therefore, the reaction system should be prepared on ice throughout the experiment, and the enzyme should be immediately stored at -20°C after use.

6. Unless otherwise specified, the cleavage reaction system described in this manual is applicable to all types of Cas12a2 enzymes provided by ToloBio.

7. This product is for research use only.

FAQ

Product Instruction

#32122 SuCas12a2 Nuclease

pdf

978.0KB

References

COA Query

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