| Products/Services Used | Details | Operation |
|---|---|---|
| Codon Optimization | For the expression of 472 Cas9 we replaced the TAL sequences of our pTALEN-pA plasmid 473 with a T7 promoter and a codon optimized Cas9 coding region 474 [35] purchased from Genscript (Piscataway, USA) to derive the 475 pCAG-Cas9-pA expression vector. | Get A Quote |
The use of TALEN and CRISPR/CAS nucleases is becoming increasingly popular as a means to edit single target sites in one-cell mouse embryos. Nevertheless, an area that has received less attention concerns the engineering of structural genome variants and the necessary religation of two distant double-strand breaks. Herein, we applied pairs of TALEN or sgRNAs and Cas9 to create deletions in the Rab38 gene. We found that the deletion of 3.2 or 9.3 kb, but not of 30 kb, occurs at a frequency of 6-37%. This is sufficient for the direct production of mutants by embryo microinjection. Therefore, deletions up to ∼10 kb can be readily achieved for modeling human disease alleles. This work represents an important s... More
