Journal
ANALYTICAL BIOCHEMISTRY
Volume 388, Issue 1, Pages 167-169Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2009.02.020
Keywords
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Funding
- NATIONAL CANCER INSTITUTE [R01CA075253] Funding Source: NIH RePORTER
- NCI NIH HHS [R01CA075253, R01 CA075253-12, R01 CA075253] Funding Source: Medline
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Preparation of heteroduplexes in large quantities with high purity is essential for the measurement of DNA mismatch repair (MMR) activity. Here we report a rapid, less labor-intensive method for the preparation of a heteroduplex plasmid that expresses the enhanced green fluorescent protein (EGFP) if the mismatch is repaired correctly. The method involves the use of a wild-type and a mutated EGFP expression plasmid and a few steps of enzymatic digestion. When the constructed heteroduplex EGFP plasmid was transfected into MMR-proficient and -deficient cell lines, the number of EGFP-expressing cells was Much higher in the MMR-proficient cells than in the MMR-deficient cells, suggesting that the heteroduplex can be used for MMR activity assay in live model systems. (C) 2009 Elsevier Inc. All rights reserved.
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