Chunk #69 — PROCEDURE — Functional testing: detection of indel mutations by the SURVEYOR nuclease assay ● TIMING 5–6 h

Source

Genome engineering using the CRISPR-Cas9 system.

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yes

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■ PAUSE POINT Extracted DNA can be stored at –20 °C for several months. 75Setting up the SURVEYOR PCR. Master-mix the following using the SURVEYOR primers provided by the CRISPR Design Tool (Step 1): ComponentAmount (μl)Final concentrationHerculase II PCR buffer, 5×101×dNTP, 100 mM (25 mM each)12 mMSURVEYOR-Fwd primer, 10 μM10.2 μMSURVEYOR-Rev primer, 10 μM10.2 μMHerculase II fusion polymerase1MgCl2, 25 mM21 mMDNA template12 ng μl−1ddH2O33Total50