T7E1 Assay Protocol

Authors

Marcia Brady

Introduction

Step1：CRISPR Delivery

Delivery methods:

Lentivirus Transduction

Plasmid transfection

Ribonucleoprotein delivery

Step2：Genome Extraction

Please follow the procedure in Genomic Extraction Kit

Or using the Direct PCR Kit to treat cell samples

Step3：Nest PCR for Target site

Nest PCR

First round PCR: 20 cycles

800-1000bp length

Second round PCR: 30-40cycles

Around 500bp length

Step4：PCR product Annealing

Step5：T7E1 treatment

Please following the procedure in Knockout Detection Kit

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

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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