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Analysis of mismatch repair in human nuclear extracts
1Oregon State University, Corvallis, OR, USA.
Abstract:
This unit describes an assay for completed mismatch correction along with a precise and sensitive assay for mismatch-repair excision intermediates and a simple and efficient procedure for construction of mismatch repair substrates. These substrates are derived by ligating synthetic oligomers into a gapped plasmid generated using the sequence-specific N.BstNBI nicking endonuclease, then the sequence-specific nicking endonuclease N.AlwI, to introduce single nicks for initiation of excision. These methods can be easily adapted for construction and analysis of DNA-lesion-containing substrates to study DNA-repair pathways other than mismatch repair.
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