Table 2. Off-target effects in bone marrow cells following intravenous treatment of β-thalassemic mice with SCF and γtcPNA4/donor DNA NPs.

Gene locus	Sequences of partial homology (5′–3′)	Size of region sequenced	Alleles sequenced	Number modified	Frequency (%)
β-globin	TGCCCTGAAAGAAAGAGA	128	8,615,313	337,192	3.9
Vascular cell adhesion protein precursor 1	AGCCCTGAAAGAAAGAGA	111	482,051	0	0
Polypyrimidine tract binding protein	GAACCTGAAAGAAAGAGA	101	355,567	2	0.00056
Protocadherin fat 4 precursor	CACCCTGAAAGAAAGAAG	115	123,158	0	0
Olfactory receptor 266	AAGCCTGAAAGAAAGATT	172	1,099,880	262	0.0231
Syntaxin binding protein	AGAAATGAAAGAAAGAGA	150	2,493,024	0	0
Muscleblind like protein	GGTGGTGAAAGAAAGAGA	165	2,336,715	0	0
Ceruloplasmin isoform	AGGACTGAAAGAAAGAGT	154	1,397,271	0	0
Total off-target			8,287,666	268	0.0032

The top seven gene loci in the mouse genome with partial homology to the 18 bp γtcPNA4 target site in β-globin intron 2 were identified, with the sequences as indicated. Thalassemic mice were treated with SCF followed by intravenous infusion with NPs containing γtcPNA4/donor DNA, and genomic DNA from c-Kit+ BM cells was subject to deep sequencing analysis at these loci. The size of the region sequenced around each site is listed, along with the number of alleles sequenced and the number of alleles with modified sequences.