Advanced Confocal Microscopy Techniques to Study Protein-protein Interactions and Kinetics at DNA Lesions

So&#328;a Legartov&#225;; Jana Such&#225;nkov&#225;; Jana Krej&#269;&#237;; Alena Kova&#345;&#237;kov&#225;; Eva B&#225;rtov&#225;

doi:10.3791/55999

Journal

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Biology

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Advanced Confocal Microscopy Techniques to Study Protein-protein Interactions and Kinetics at DNA Lesions

JoVE Journal
Biology

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JoVE Journal Biology

Advanced Confocal Microscopy Techniques to Study Protein-protein Interactions and Kinetics at DNA Lesions

10,890 Views

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12:43 min

•

November 12, 2017

DOI:

10.3791/55999-v

DOI:

10.3791/55999-v

•

12:43 min

November 12, 2017

•

1 Views

Soňa Legartová*¹, Jana Suchánková*¹, Jana Krejčí¹, Alena Kovaříková¹, Eva Bártová¹

¹Institute of Biophysics of the Czech Academy of Sciences

Transcript

The overall goal of these experiments is to induce DNA lesions using local micro irradiation of cells and to observe the recruitment of proteins involved in DNA repair processes.This method can help us to understand a very complex and profound question such as, how normal and cancer cells respond to DNA injury.The main advantage of this technique is that we are able to study the DNA damage response in individual live cells.Demonstrating the procedure will be by Sona Legartova, a post-doc and Jana Suchankova, a PhD student from my laboratory.To begin, cultivate HeLa cells in DMAM supplemented with 10%FBS and appropriate antibiotics.Incubate the cells at 37

Summary

Laser microirradiation is a useful tool for studies of DNA repair in living cells. A methodological approach for the use of UVA lasers to induce various DNA lesions is shown. We have optimized a method for local microirradiation that maintains the normal cell cycle; thus, irradiated cells proceed through mitosis.