New in vitro dna polymerase activity and fidelity assay reveals age-dependent changes in Arabidopsis Thaliana

Andrey Golubov; Priti Maheshwari; Andriy Bilichak; Igor Kovalchuk

doi:10.4081/pb.2011.e7

New in vitro dna polymerase activity and fidelity assay reveals age-dependent changes in Arabidopsis Thaliana

https://doi.org/10.4081/pb.2011.e7

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Vol. 2 No. 1 (2011)

Submitted: 4 March 2011
Accepted: 6 June 2011

Published: 20 June 2011

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Authors

Andrey Golubov
Priti Maheshwari
Andriy Bilichak
Igor Kovalchuk
igor.kovalchuk@uleth.ca
University of Lethbridge, Canada.

DNA polymerase is an enzyme that adds nucleotides to the growing DNA chain during replication and DNA repair. DNA polymerase activity and fidelity are important characteristics that reflect the ability of DNA polymerase to add nucleotides and then proofread newly synthesized DNA. We have developed a protocol allowing analysis of polymerase activity and fidelity using crude Arabidopsis thaliana plant extracts. It is based on the ability of DNA polymerases in the extract to elongate the fluorescently labelled primer annealed to a short complementary template. For analysis, fluorescently labelled products were separated on a denaturing polyacrylamide gel and visualized using a high performance blot imager. Analysis of tissue prepared from 5-, 12- and 21-day-old Arabidopsis plants showed an age-dependent decrease in polymerase activity, an increase in polymerase fidelity and a tendency to an increase in exo- (endo) nucleolytic activity.

Igor Kovalchuk, University of Lethbridge

Professor, Department of Biological Sciences

Ethics Approval

Supporting Agencies

Golubov, A., Maheshwari, P., Bilichak, A., & Kovalchuk, I. (2011). New in vitro dna polymerase activity and fidelity assay reveals age-dependent changes in Arabidopsis Thaliana. International Journal of Plant Biology, 2(1), e7. https://doi.org/10.4081/pb.2011.e7