Plant Gateway 3-Fragment MultisitePro CRISPR-Cas9 design

This cloning strategy allows for the simultaneous expression of two guide RNAs driven by the U6 promoter (pEN-Comaira.1, pEN-Comaira.2) as well as Cas9 (pEN-RC9.3) driven by the RPS5a promoter.

The binary vector that we have chosen to use in conjunction is pEARLEYGATE-302v2, which has both KanR and AmpR selection in bacteria and Basta-selection in plants.