U6 promoter U6 promoter(1-249) U6 pr... U6 promoter(1-249) U6 pr... U6 promoter(1-249) gRNA scaffold gRNA scaffold(268-343) gRNA ... gRNA scaffold(268-343) gRNA ... gRNA scaffold(268-343) CMV enhancer CMV enhancer(440-725) CMV e... CMV enhancer(440-725) CMV e... CMV enhancer(440-725) hybrid intron hybrid intron(1005-1232) hybri... hybrid intron(1005-1232) hybri... hybrid intron(1005-1232) Kozak sequence Kozak sequence(1244-1253) Kozak... Kozak sequence(1244-1253) Kozak... Kozak sequence(1244-1253) 3xFLAG 3xFLAG(1253-1318) 3xFLA... 3xFLAG(1253-1318) 3xFLA... 3xFLAG(1253-1318) SV40 NLS SV40 NLS(1325-1345) SV40 ... SV40 NLS(1325-1345) SV40 ... SV40 NLS(1325-1345) Cas9 Cas9(1370-5470) Cas9 Cas9(1370-5470) nucleoplasmin NLS nucleoplasmin NLS(5471-5518) nucle... nucleoplasmin NLS(5471-5518) ...nucle nucleoplasmin NLS(5471-5518) T2A T2A(5534-5587) T2A T2A(5534-5587) T2A T2A(5534-5587) PuroR PuroR(5588-6187) PuroR PuroR(5588-6187) bGH poly(A) signal bGH poly(A) signal(6221-6428) bGH p... bGH poly(A) signal(6221-6428) ...bGH p bGH poly(A) signal(6221-6428) AAV2 ITR AAV2 ITR(6437-6566) AAV2 ... AAV2 ITR(6437-6566) ...AAV2 AAV2 ITR(6437-6566) AAV2 ITR AAV2 ITR(6437-6577) AAV2 ... AAV2 ITR(6437-6577) ...AAV2 AAV2 ITR(6437-6577) f1 ori f1 ori(6652-7107) f1 or... f1 ori(6652-7107) AmpR promoter AmpR promoter(7389-7493) AmpR ... AmpR promoter(7389-7493) ...AmpR AmpR promoter(7389-7493) AmpR AmpR(7494-8354) AmpR AmpR(7494-8354) ori ori(8525-9113) ori ori(8525-9113) nucleoplasmin NLS nucleoplasmin NLS(5473-5520) nucle... nucleoplasmin NLS(5473-5520) ...nucle nucleoplasmin NLS(5473-5520) gRNA scaffold gRNA scaffold(270-345) gRNA ... gRNA scaffold(270-345) gRNA ... gRNA scaffold(270-345) AmpR promoter AmpR promoter(7391-7495) AmpR ... AmpR promoter(7391-7495) ...AmpR AmpR promoter(7391-7495) AAV2 ITR AAV2 ITR(6439-6579) AAV2 ... AAV2 ITR(6439-6579) ...AAV2 AAV2 ITR(6439-6579) AmpR AmpR(7496-8356) AmpR AmpR(7496-8356)