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Data sheet
| Size | 50 µg |
| Background | In molecular biology, CRISPR-associated endonuclease in Prevotella and Francisella 1 or Cpf1 is a single RNA-guided endonuclease lacking a small trans-encoded RNA, a tracrRNA, and it utilizes a T-rich protospacer-adjacent motif of 2-6 base pair DNA sequence immediately following the DNA sequence targeted by the Cas9 nuclease in the CRISPR bacterial adaptive immune system, also known as PAM. It recognizes a T-rich PAM, TTTN, but on the 5' side of the guide. Cpf1 cleaves DNA via a staggered DNA double-stranded break. Novateinbio Lb Cas12a (Cpf1) is a programmable DNA endonuclease guided by a single guide RNA (gRNA). Targeting requires a gRNA complementary to the target site as well as a 5' TTTN protospacer adjacent motif (PAM) on the DNA strand opposite the target sequence. Cleavage by Novateinbio Lb Cas12a (Cpf1) occurs ~18 bases 3' of the PAM and leaves 5' overhanging ends. |
| Host/Source | E. coli |
| Physical Appearence | Sterile filtered colorless solution. Or Powder |
| Concentration | 50 μg/ 50 μl (260 pmol, 5.2 uM ) |
| Storage and Stability | ‐70°C. Avoid freeze thaw cycles. |
More info
Cas9 vs. Cas12a
Cas9 requires two RNA molecules to cut DNA while Cas12a needs one. The proteins also cut DNA at different places, offering researchers more options when selecting an editing site. Cas9 cuts both strands in a DNA molecule at the same position, leaving behind blunt ends. Cas12a leaves one strand longer than the other, creating sticky ends. The sticky ends have different properties than blunt ends during non-homologous end joining or homologous repair of DNA, which confers certain advantages to Cas12a when attempting gene insertions, compared to Cas9. Although the CRISPR/Cas9 system can efficiently disable genes, it is challenging to insert genes or generate a knock-in. Cas12a lacks tracrRNA, utilizes a T-rich PAM and cleaves DNA via a staggered DNA DSB.
In summary, important differences between Cas12a and Cas9 systems are that Cas12a:
• Recognizes different PAMs, enabling new targeting possibilities.
• Creates 4-5 nt long sticky ends, instead of blunt ends produced by Cas9, enhancing the efficiency of genetic insertions and specificity during NHEJ or HDR.
• Cuts target DNA further away from PAM, further away from the Cas9 cutting site, enabling new possibilities for cleaving the DNA.
|
Feature |
Cas9 |
Cas12a |
|
Structure |
Two RNA required (Or 1 fusion transcript (crRNA+tracrRNA=gRNA) |
One crRNA required |
|
Cutting mechanism |
Blunt end cuts |
Staggered end cuts |
|
Cutting site |
Proximal to recognition site |
Distal from recognition site |
|
Target sites |
G-rich PAM |
T-rich PAM |
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