We are working with E.coli K12 strain and want to knock out a certain gene in its genome. We already have some concepts of sgRNA, but we do not know in which plasmid (either pCas9 or pGRB or pCRISPR) we should ligate it. Should the Cas9 AND sgRNA be expressed both in the same vector, or sepperate? and is a certain promotor for the expression for sgRNA a must (e.g. the U6 promotor)?

Thanks in advance!