Make your target specific gRNA lentivectors with desired markers!
This Cloning Kit is designed for cloning of double-strand DNA encoding your desired gRNA sequence for CRISPR genomic gene editing.
To make your target specific gRNA lentivector, two synthetic gRNA oligonucleotides are first annealed to form the double stand DNA duplex which is then cloned into the provided, linear lentivector via T4 enzyme ligation.
1) Flexibility to select desired promoter: H1 or U6;
2) Flexibility to choose desired selection marker: antibiotic or antibiotic-fluorescent fusion dual marker;
This product is the lentivector version having the gRNA driven by a constitutive human U6 promoter with a GFP-Blastidicin dual selection marker under Rsv promoter. Please see Product Manual for details.
pLenti-U6-(GFP-Bsd) linear vector: 10 ul (10rxn, 1ul/rxn)
10x gRNA oligo annealing solution: 25 ul
10X ligation buffer: 20 ul
T4 ligase enzyme: 10 ul (10rxn, 1ul/rxn)
Cloning control insert: annealed Neg-gRNA duplex: 10 ul (10rxn, 1ul/rxn)
Sequencing primer: 10 ul (10xn, 1ul/rxn)
DH5a chemical competent cells: 10 vials (10xn, 1 vial/rxn)
The generated gRNA lentivector can be used for transfection in CRISPR editing, or used for gRNA lentivirus production and then use the gRNA lentivirus for CRISPR editing.