Genomic DNA from cell pellets was extracted using QuickExtract DNA Extraction Solution 1.0 (Epicentre). Amplification of the junction regions surrounding gene KI sites in edited cells was carried out by PCR using Q5 High-Fidelity DNA polymerase (New England BioLabs) and 100 ng genomic DNA as template in a 50 μL reaction. PCR products were purified using QlAquick PCR purification kit (Qiagen) and then subjected to direct DNA sequencing service (ACGT, Wheeling, IL). The indel rates were analyzed by the online software12 (link) (http://tide.nki.nl ) using sequence without indels as reference.
Protocol detail
Quantifying Gene Editing Efficiency
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Corresponding Organization :
Other organizations : University of Illinois Urbana-Champaign
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Variable analysis
independent variables
- Gene KI sites in edited cells
- Indel rates
- Genomic DNA from cell pellets
- 100 ng genomic DNA as template
- Sequence without indels as reference
- Positive control: None specified
- Negative control: None specified
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