Table 1.

Optimization of short guide RNA targeting murine Drd1a gene Cel-I assay and off-target prediction in silico

No.	Primers	Sequence	Strand	Domain	PAM	Cleavage efficiency (%)	Off target score (predicted)
1	Forward	5′-CACCGATGGCTCCTAACACTTCTACC-3′	–	N-term	ATGGAT	9.4	83.9
	Reverse	5′-AAACGGTAGAAGTGTTAGGAGCCATC-3′
2	Forward	5′-CACCGAGATTGCTGGCTTTTGGCCCT-3′	–	TM2	TTGGGT	5.2	77.5
	Reverse	5′-AAACAGGGCCAAAAGCCAGCAATCTC-3′
3	Forward	5′-CACCGCATAAATAATGGGGTTCAGGG-3′	+	TM7	AGGAAT	0.0	59.9
	Reverse	5′-AAACCCCTGAACCCCATTATTTATGC-3′
4	Forward	5′-CACCGCCTGATGACAGCGGCACAGAC-3′	+	TM1	AAGGGT	12.0	78.0
	Reverse	5′-AAACGTCTGTGCCGCTGTCATCAGGC-3′
5	Forward	5′-CACCGACAGACAAGGGTATTCCCTAA-3′	+	TM2	GAGAGT	6.0	87.6
	Reverse	5′-AAACTTAGGGAATACCCTTGTCTGTC-3′
6	Forward	5′-CACCGCTTATCCTGTCCACTCTCTTA-3′	–	TM1	GGGAAT	22.1	73.3
	Reverse	5′-AAACTAAGAGAGTGGACAGGATAAGC-3′
7	Forward	5′-CACCGGGTATTCCCTAAGAGAGTGGA-3′	+	TM2	CAGGAT	11.7	73.0
	Reverse	5′-AAACTCCACTCTCTTAGGGAATACCC-3′
8	Forward	5′-CACCGCAGGGACAGAAAACAGGCTGT-3′	+	TM1	GAGGAT	23.7	48.4
	Reverse	5′-AAACACAGCCTGTTTTCTGTCCCTGC-3′