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T7E1 Assay Protocol

t7e1 assay protocol 01Step1:CRISPR Delivery

Delivery methods:

Lentivirus Transduction

Plasmid transfection

Ribonucleoprotein delivery

t7e1 assay protocol 02Step2:Genome Extraction

Please following the procedure in Genomic Extraction Kit

Or using the Direct PCR Kit to treat cell samples

t7e1 assay protocol 03 Step3:Nest PCR for Target site

Nest PCR

First round PCR: 20 cycles

800-1000bp length

Second round PCR: 30-40cycles

Around 500bp length

t7e1 assay protocol 04 Step4:PCR product Annealing

Annealing procedure

t7e1 assay protocol 07

t7e1 assay protocol 05 Step5:T7E1 treatment

Please following the procedure in Knockout Detection Kit

Or using T7E1 enzyme for incubation in 37℃ for 30min

t7e1 assay protocol 06 Step6:Gel electrophoresis

Data shall be visualized by 1.2%-1.5% agarose gel

Note: direct PCR product could be used for T7E1 detection.

After treated with T7E1, reactions should be electrophoresed by high concentration agarose gels.

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