HomeFAQsWhy do I observe incomplete digestion with EnGen® Sau Cas9 and my target-specific Sau sgRNA in vitro?
FAQ: Why do I observe incomplete digestion with EnGen® Sau Cas9 and my target-specific Sau sgRNA in vitro?
The efficiencies of sgRNAs vary depending on the specific target sequences. We recommend using an on-line tool (which use certain factors such as GC content) to select the best target site within your DNA of interest. We also recommend purification of the dsDNA target and verifying the integrity of the sgRNA on a denaturing gel. If sgRNA efficiency is low, increasing the molar ratio of sgRNA:Sau Cas9:target DNA to 10:10:1 or higher may improve DNA cleavage. We also recommend using the supplied NEBuffer™ 3.1 for best results.