Experimental design considerations for efficient and specific gene knockin using a CRISPR-Cas9 for HDR with synthetic crRNA and tracrRNA
Poster Aug 19, 2016
Hidevaldo B. Machado, John A. Schiel, Maren Mayer-Gross, Eldon T. Chou, Melissa L. Kelley, Anja van Brabant Smith. Dharmacon, part of GE Healthcare, 2650 Crescent Drive, Lafayette, CO 80026, USA
The bacterial CRISPR-Cas9 system has revolutionized the genome engineering world with its efficiency and ease of use. The most common use of the CRISPR-Cas9 technology has been to engender gene knockouts that are generated as a result of imperfect repair of a targeted double-strand DNA break by the non-homologous end joining (NHEJ) pathway.
When there is a need to quickly analyze samples using a number of different PCR assays, it is likely that optimal conditions for each assay will not be the same. First, different assays often will require different annealing temperatures for their primers. In addition, amplicons may be designed to be of different lengths and therefore require varying durations of the extension step.READ MORE