GenCRISPR™ ゲノム編集関連製品 » GenCRISPR™ ヌクレアーゼ-CRISPR遺伝子編集 » GenCRISPR™ LbuCas13a Nuclease
GenCRISPR™ LbuCas13a Nuclease

LbuCas13a has sequence-specific RNA cleavage activity, and after specific cleavage, it activates collateral cleavage activity towards adjacent non-specific ssRNA sequences.

A 50 μL reaction containing ssRNA, Cas13a/crRNA complex, Fluorescent RNA reporter, and Cas13a Reaction Buffer is incubated for 30 minutes at 37 °C. After incubation, the fluorescence intensity is measured using a Microplate Reader (Excitation wavelength: 494 nm; Emission wavelength: 518 nm).

GenCRISPR™ LbuCas13a Nuclease

GenCRISPR™ LbuCas13a Nuclease is an RNA-guided, RNA-targeting CRISPR enzyme, commonly referred to as LbuC2c2. This Cas13a protein can be applied to detect single RNA molecules with proven high specificity. Recently, Cas13a has been leveraged as a diagnostic nucleic acid detection tool.
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Overview
Description GenCRISPR™ LbuCas13a Nuclease is an RNA-guided RNA endonuclease from Leptotrichia buccalis. Cas13a (previously known as C2c2) belongs to the Class 2 Type VI CRISPR-Cas system and contains a single protein effector with two HEPN domains. Cas13a exhibits two distinct ribonuclease activities. Cas13a possesses sequence-specific RNA cleavage activity in vitro and in vivo. And after specific cleavage, Cas13a activates collateral cleavage activity towards adjacent non-specific ssRNA sequences. The two distinct ribonuclease activities of Cas13a enable guide-RNA processing and RNA detection. The RNA detection empowered with Cas13a-crRNA complex and quenched fluorescent ssRNA as reporter has been reported to be versatile, rapid and high-specific. 

Applications
crRNA-dependent RNA cleavage

Properties
Source Recombinant Cas13a expressed by E.coli
Species Leptotrichia buccalis
Tag Tag-free
Apparent Molecular Weight ~140 kDa, on SDS-PAGE under non-reducing conditions
Concentration Please refer to the COA for the specific lot.
Storage Buffer 10 mM Tris-HCl, 300 mM NaCl, 1.0 mM DTT, 50% glycerol, pH 8.0. 
Storage & Stability Store at -20 °C for up to 12 months from the date of manufacture. Avoid repeated freeze-thaw cycles. Do not store below -20 °C!
Accession# C7NBY4

Quality Control Specifications
Appearance Clear, colorless liquid
Purity ≥ 90% by SDS-PAGE
Activity ≥ 85% by in vitro digestion of ssRNA
Concentration 4 mg/ml ± 10% as analyzed by A280
Residual DNase Undetectable
Residual RNase Undetectable
Endotoxin Level < 0.2 EU/μg

Examples
  • GenCRISPR™ LbuCas13a Nuclease
    • GenCRISPR™ LbuCas13a Nuclease

    LbuCas13a has sequence-specific RNA cleavage activity, and after specific cleavage, it activates collateral cleavage activity towards adjacent non-specific ssRNA sequences.

    A 50 μL reaction containing ssRNA, Cas13a/crRNA complex, Fluorescent RNA reporter, and Cas13a Reaction Buffer is incubated for 30 minutes at 37 °C. After incubation, the fluorescence intensity is measured using a Microplate Reader (Excitation wavelength: 494 nm; Emission wavelength: 518 nm).


Background
References 1.       Abudayyeh, Omar O., et al. "C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector." Science 353.6299 (2016).
2.       Gootenberg, Jonathan S., et al. "Nucleic acid detection with CRISPR-Cas13a/C2c2." Science 356.6336 (2017): 438-442.
3.       Gootenberg, Jonathan S., et al. "Multiplexed and portable nucleic acid detection platform with Cas13, Cas12a, and Csm6." Science 360.6387 (2018): 439-444.
4.       Iwasaki, Roman S., and Robert T. Batey. "SPRINT: a Cas13a-based platform for detection of small molecules." Nucleic acids research 48.17 (2020): e101-e101.
5.       East-Seletsky, Alexandra, et al. "Two distinct RNase activities of CRISPR-C2c2 enable guide-RNA processing and RNA detection." Nature 538.7624 (2016): 270-273.

For laboratory research use only. Direct human use, including taking orally and injection and clinical use are forbidden.