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− | <p>For this part, we have deleted the HNH domain, Rec2 domain and RuvCIII-2 domain of the SP-dCas9. The triple truncation (∆REC2 ∆RuvCIII-2 ∆HNH, referred to as ‘∆3ple’) is especially of interest | + | <p>For this part, we have deleted the HNH domain, Rec2 domain and RuvCIII-2 domain of the SP-dCas9. The triple truncation (∆REC2 ∆RuvCIII-2 ∆HNH, referred to as ‘∆3ple’) is especially of interest since it reduces the size of the dCas9 gene to around 3.2kB which is on par with the size of Sa-Cas9. The ∆3ple truncation has slightly lower activation compared to double truncation. In order to improve the activity of ∆3ple dCas9, quick changes are further performed at the various site of dCas9. Combination of quick change at Asn497Lys (QC5), Thr657Lys (QC6) and Ser1109Arg (QC15) show improved activity. Its binding capability was tested by assessing the strength in transcriptional activation via by tagging it with a VP64-p65-Rta tripartite activator.</p> |
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Revision as of 19:17, 1 November 2017