After drug treatments I would like to check a number of genes by qPCR to determine involvement of different DNA damage pathways namely homologous recombination (HR), NHEJ, MMEJ etc. Which genes will be best to check to get a quick idea?
For DNA double strand break, you can use gamma H2AX.
Article ??-H2AX- A novel biomaker for DNA double-strand breaks
gH2AX is not a universal marker and definitely not the gene to determine DDR. if talking about DNA breaks - you may want to focus of some Fra sites. If you want to see specific DDR pathway involved, you would need specific system for each type of repair as they are uncoupled in most of the systems. Am attaching Amanda Gunn and Jeremy M. Stark paper with detailed protocol.
You could try doing western blot analysis better. For example, phosphor-RPA detection is a well-known mark of DNA end resection, that occurs in HR pathways. On the other hand, phospho-DNAPKcs (T2609) might be a good marker for NHEJ
Thank you Lyakhovich and Ruiz for your suggestions. I will check a number of markers at protein level. However, for qPCR I am thinking to use commercially available gene set for different DNA damage. Thanks again.
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