先导编辑系统(Prime editor,PE) plasmid质粒载体 BioVector NTCC质粒载体菌种细胞基因保藏中心

先导编辑系统(Prime editor,PE) plasmid质粒载体

The PE system retains CRISPR’s targeting specificity but carries with it, additional cargo in the form of an edit-containing RNA template as a contiguous extension of the guide RNA (known as the pegRNA), and M-MLV reverse transcriptase (RT) fused to the C terminus of Cas9 (H840A) nickase. Use of the Cas9 nickase avoids the formation of a DSB, and simply cuts the non-complementary strand of the DNA three bases upstream of the PAM site. This exposes a DNA flap with a 3’ OH group which binds to the primer binding site (PBS) of the RNA template, serving as a primer for RT, which extends the 3’ flap by copying the edit sequence of the pegRNA (Fig. 1). Despite this extended 3’ flap being thermodynamically less likely to hybridise to the unedited complementary strand compared to the unedited 5’ flap, the inherent preference of the endogenous endonuclease FEN1 to excise 5’ flaps leads to hybridisation of the edited 3’ flap being favoured, thus resulting in highly efficient base editing. And this was PRIME version 1.0.

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