. 2019 Jul 29;39(16):e00105-19. doi: 10.1128/MCB.00105-19

TABLE 1.

Characterization of the U2OS RB1 mutant clones generated by targeting exon 22^a

Clone	Cas9 form	RB1 genotype^b	Predicted protein^c	Off-target genotype^d
Clone	Cas9 form	RB1 genotype^b	Predicted protein^c	ALDH1L1	ZNF699
U2OS	NA	Wild type	WT	NA	NA
U2OS	NA	Wild type	WT	NA	NA
21B2	D10A	Wild type	WT	ND	ND
21B2	D10A	Wild type	WT	ND	ND
21B4	D10A	Wild type	WT	WT	WT
21B4	D10A	Wild type	WT	WT	WT
11B5	D10A	Wild type	WT	ND	ND
11B5	D10A	Wild type	WT	ND	ND
4C1	D10A	Heterozygous	WT	WT	WT
4C1	D10A	Heterozygous	p.Y771fsX8	WT	WT
21D5	D10A	Heterozygous	WT	WT	WT
21D5	D10A	Heterozygous	p.Q770HdelX10	WT	WT
21A1	D10A	Heterozygous	WT	WT	WT
21A1	D10A	Heterozygous	p.I753delX12	WT	WT
12C3	D10A	Null	p.I752IfsX9	WT	WT
12C3	D10A	Null	p.E748CdelX11	WT	WT
5A5	WT	Null	p.N757TX31	WT	WT
5A5	WT	Null	p.P776VdelX6	WT	WT
6B1	WT	Null	p.Y756YfsX14	WT	WT
6B1	WT	Null	p.S751SfsX10	WT	WT
5C4	WT	Null	p.S773FfsX14	WT	WT
5C4	WT	Null	p.Y756delX	WT	WT

NA, not applicable; ND, not determined; WT, wild type.

Genotypes were determined by PCR and sequencing.

Amino acid coding changes were predicted based on nucleotide sequences.

The top-scoring off-target intragenic locations determined for each gRNA, ZNF699 for gRNA X22B and ALDH1L1 for gRNA X22C, were also sequenced to probe for unwanted mutations.