Methods and compositions for alteration of a cystic fibrosis transmembrane conductance regulator (CFTR) gene

摘要

Nucleases and methods of using these nucleases for alteration of a CFTR gene and generation of cells and animal models.

主权项

1. A protein comprising an engineered zinc finger protein DNA-binding domain, wherein the DNA-binding domain comprises four, five or six zinc finger recognition regions ordered F1 to F4, F1 to F5 or F1 to F6 from N-terminus to C-terminus, and wherein the protein is selected from the group consisting of:
(i) a 5-finger zinc finger protein comprising the following recognition regions:
F1: WPSCLYA (SEQ ID NO:8);F2: NGVLLKR (SEQ ID NO:9);F3: QSGNLAR (SEQ ID NO:10);F4: RSDNLSE (SEQ ID NO:11); andF5: NPRNRFT (SEQ ID NO:12), wherein the protein binds to a target site as shown in SEQ ID NO:37; (ii) a 5-finger zinc finger protein comprising the following recognition regions:
F1: RSDVLSE (SEQ ID NO:13);F2: QSGNLAR (SEQ ID NO:10);F3: QSGHLSR (SEQ ID NO:14);F4: RSDVLSE (SEQ ID NO:15); andF5: WSASLSK (SEQ ID NO:16), wherein the protein binds to a target site as shown in SEQ ID NO:38; (iii) a 4-finger zinc finger protein comprising the following recognition regions:
F1: QNATRIN (SEQ ID NO:17);F2: QSGNLAR (SEQ ID NO:10);F3: RSDNLST (SEQ ID NO:18); andF4: QSADRKK (SEQ ID NO:19), wherein the protein binds to a target site as shown in SEQ ID NO:39; (iv) a 6-finger zinc finger protein comprising the following recognition regions:
F1: TNQNRIT (SEQ ID NO:20);F2: RNQTRIT (SEQ ID NO:21);F3: QSGNLAR (SEQ ID NO:10);F4: QSNTRIM (SEQ ID NO:22);F5: TSGNLTR (SEQ ID NO:23); andF6: QSNALHQ (SEQ ID NO:24), wherein the protein binds to a target site as shown in SEQ ID NO:40; (v) a 5-finger zinc finger protein comprising the following recognition regions:
F1: TSSDRKK (SEQ ID NO:25);F2: QSSDLSR (SEQ ID NO:26);F3: DRSNLTR (SEQ ID NO:27);F4: TSGNLTR (SEQ ID NO:23); andF5: WRLSLQV (SEQ ID NO:28), wherein the protein binds to a target site as shown in SEQ ID NO:41; (vi) a 6-finger zinc finger protein comprising the following recognition regions:
F1: QSGNLAR (SEQ ID NO:10);F2: QGANLIK (SEQ ID NO:29);F3: RSDHLSA (SEQ ID NO:30);F4: ESRYLMV (SEQ ID NO:31);F5: RSDNLST (SEQ ID NO:18); andF6: DRSNRKT (SEQ ID NO:32), wherein the protein binds to a target site as shown in SEQ ID NO:42; (vii) a 5-finger zinc finger protein comprising the following recognition regions:
F1: TSGNLTR (SEQ ID NO:23);F2: QSNALHQ (SEQ ID NO:24);F3: QSGNLAR (SEQ ID NO:10);F4: TSGNLTR (SEQ ID NO:23); andF5: WWTSRAL (SEQ ID NO:33), wherein the protein binds to a target site as shown in SEQ ID NO:43; and (viii) a 4-finger zinc finger protein comprising the following recognition regions:
F1: HSNARKT (SEQ ID NO:34);F2: TSGNLTR (SEQ ID NO:23);F3: TLQNRMS (SEQ ID NO:35); andF4: DQSTLRN (SEQ ID NO:36), wherein the protein binds to a target site as shown in SEQ ID NO:44.