Fluorescence imaging to understand the molecular mechanism of DNA damage-triggered cellular reprogramming in plants

• Main Authors: Tamada, Yosuke, Imai, Akihiro, Gu, Nan
• Format: Conference Proceeding
• Language: English
• Published: SPIE 27.10.2021
• ISBN: 1510647198; 9781510647190
• ISSN: 0277-786X
• DOI: 10.1117/12.2615404

We recently found that massive but transient DNA strand breaks, a kind of DNA damage, can trigger cellular reprogramming of differentiated cells to stem cells in the moss Physcomitrella patens (Physcomitrella). In Physcomitrella, DNA is massively damaged by the treatment with a DNA strand break-inducing reagent at high concentration. The DNA damage was repaired within 24 hours after the removal of the reagent. Then, the promoter activity of STEMIN1, encoding an integrator of the reprogramming signals, became active in a part of leaf cells, which eventually became stem cells. Levels of DNA strand breaks were quantified by performing comet assay, where the nuclei are subject to electrophoresis, stained with the fluorescent dye that binds to DNA, and observed with fluorescent microscope. The promoter activity of STEMIN1 was visualized using fluorescent protein and observed with fluorescent microscopy.