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Pipetting scheme for sgRNA amplification PCR.
| Component | Amount per reaction (µl) | Final concentration |
|---|---|---|
| NEBNext High Fidelity PCR Master Mix, 2 × | 25 | 1x |
| Pooled sgRNA library template | 10.4 ng/µl | |
| NGS-Lib-Fwd primer (unique), see Table 9 | 1.25 | 0.25 µM |
| NGS-Lib-KO-Rev, see Table 9 | 1.25 | 0.25 µM |
| Pyrogen/RNase/DNase free water | add to a total volume of 50 µl |
