Part:BBa_K1723021

dCas9_VP64

Cas9 (CRISPR associated protein 9) is an RNA-guided DNA endonuclease that targets and cleaves any DNA sequence complementary to its guide RNA (gRNA). Catalytically “dead” Cas9 (dCas9) lacks the ability to cleave DNA. Fused to both SV40 nuclear localization sequence and VP64 eukaryotic transcription activation domain, chimeric dCas9_VP64 can act as a programmable transcription activator or inhibitor when forming a complex with the appropriate guide RNA (gRNA). The gRNA exhibits a 20 base pair specificity determinant sequence (SDS). The gRNA-dCas9_VP64 complex will bind to a similar SDS in the genome situated next to a consensus PAM sequence (NGG). Activation occurs by binding of a gRNA-dCas9_VP64 complex upstream of the promoter, thus recruiting RNA Polymerase II via VP64. Inhibition occurs by binding of a gRNA-dCas9_VP64 complex on the promoter, thus sterically hindering the binding of RNA Polymerase II on the promoter [1].

To learn more about how this part was used specific to our project, please follow this link: http://2015.igem.org/Team:EPF_Lausanne/Part_Collection

Usage and Biology

S. cerevisiae