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Homology-directed gene repair. exogenous repair substrate. I-SceI. xx. sister chromatids. xx. Rad51…. Sequence correction. xx. Homology-directed gene repair. exogenous repair substrate. I-SceI. xx. sister chromatids. xx. Rad51…. Sequence correction. xx. DRGFP reporter.
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Homology-directed gene repair exogenous repair substrate I-SceI xx sister chromatids xx Rad51… Sequence correction xx
Homology-directed gene repair exogenous repair substrate I-SceI xx sister chromatids xx Rad51… Sequence correction xx
DRGFP reporter CA puroR I-SceI STOP BcgI gene conversion CA GFP+ puroR BcgI BcgI
041408 293T stable DRGFP (S8) pCMV-I-SceI no DNA 48 hrs 72 hrs
041408 293T DRGFP (cis) DRGFP+I-SceI I-SceI DRGFP
041408 293T DRGFP-lacO (cis) DRGFPlacO+I-SceI DRGFPlacO
041408 293T ZFGFP + iGFP (trans) ZFGFP+iGFP ZFGFP+iGFP+I-SceI
041408 293T ZFGFP-lacO + iGFP (trans) ZFGFP-lacO ZFGFP-lacO+iGFP+I-SceI
050208 293T: Efficiency of Transient Transfection no DNA 62 ng eGFP 125 ng eGFP 250 ng eGFP 500 ng eGFP 1.0 g eGFP 2.0 g eGFP 4.0 g eGFP
“Stable” DRGFP (puroS) 48 hrs 73 hrs 98 hrs No DNA CMV-Sce
“Stable” DRGFP (puroR) 48 hrs 73 hrs 98 hrs No DNA CMV-Sce
Plans • Lines with integrated trans reporter • Enhance repair by tethering chromatin modifiers and repair factors • Back to B-cells