Our months of lab work contributed to the construction of several new parts. Please consult the registry for a more detailed explanation of our parts and the validation work that was performed for it.

Best Basic Part: dCas9 VRER

Our favorite part and the Groningen contender for Best New Basic part is dCas9 VRER. This variation on our improved spdCas9s for bacterial use, is adapted in such a way that it recognizes different PAM sites. These different sites ensure that the two Cas-complexes of our system do not compete for the binding sites of their counterpart. The difference in PAM sites ensures that the dCas9 VRER protein only binds to the sites which are specifically located on our pre-programmed reporter plasmid. It also prevent it from binding on the injected phage DNA. This also works the other way around as hCas9 only binds on the infecting phage DNA and not on the reporter plasmid, as it would cleave the plasmid upon binding to one of its sites.

Parts list
.gb file Part NumberTypePart NameAuthorbp
BBa_K2361001CodingdCas9 VRERMart Bartelds4108
  BBa_K2361000CodingspdCas9Mart Bartelds4120
  BBa_K2361003RNApUSP45 tracrRNAThijs Ettema474
  BBa_K2361004RNACRISPR arraySebald Verkuijl639
  BBa_K2361005RNACRISPR array containing a 20 spacerSebald Verkuijl691
  BBa_K2361006RNACRISPR array containing a 21 spacerSebald Verkuijl691
  BBa_K2361007RNACRISPR array containing empty spacersSebald Verkuijl625
  BBa_K2361009RegulatoryNisin inducable lactococcal promoterThijs Ettema198
BBa_K2361002CodinghCas9 operonThijs Ettema6060
BBa_K2361008Regulatoryp32 lactococcal promoterThijs Ettema196
BBa_K2361010RegulatorypUSP45 lactococcal promoterThijs Ettema606

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