DNA mismatch repair detected in human cell extracts.

摘要: A system to study mismatch repair in vitro HeLa cell extracts was developed. Preformed heteroduplex plasmid DNA containing two single base pair mismatches within the SupF gene of Escherichia coli used as a substrate assay. Repair one or both wild-type sequence measured by transformation lac(Am) E. strain which presence an active supF could be scored. The constructed carry mutations genes associated with and recombination (mutH, mutU, recA) so that processing bacterium minimal. Extract reactions were carried out incubation ATP, creatine phosphate, kinase, deoxynucleotides, magnesium-containing buffer added. Under these conditions about 1% repaired. In absence added energy sources activity significantly reduced. addition either aphidicolin dideoxynucleotides reduced activity, but only effective blocking polymerization extracts. It is concluded energy-requiring process dependent on adequate deoxynucleotide concentration. results also indicate some type polymerization, different effects suggest cannot simply accounted for random nick-translation alone.