We designed FRET-FISH probes for optimization of the design with the first being on MYC gene on human HAP1 cell lines and Ogt on mouse NIH3T3 and MEF.
Thereafter, the optimized FRET-FISH design targeted Magix, Kdm5c, Atp2b3, Tent5d, Ddx3x and Pbd1c on Chr X and Atp5a1, Hspa9, Nars, Minar2, Grxcr2 and 4930426D05Rik on Chr 18. A FRET-FISH probe spans around 100 kb. The experiments were replicated.
Experiment Imaging Method: epifluorescence microscopy
Protocol Name: FRET-FISH protocol
Each dataset consists of multiple fields of view (FOVs) that were acquired with 2 different dyes and the FRET channel that consists using the excitation source of the donor dye and filter the fluorescence signal with the emission filters of the acceptor dye. (AF488 labeled as ‘a488’; Cy3 as ‘tmr’; AF594 as ‘a594’; Cy5 as ‘Cy5’ and Hoechst 33342 as ‘dapi’) and all the FOVs are provided as separate .tiff files. Each FOV comprises 40–50 focal planes spaced 0.3 μm apart and has a size of 1024x1024 pixels.
Swedish Cancer Society (grant no. CAN 2018/728 and grant no. 19 0130 Pj 03 H)
Science for Life Laboratory, the Karolinska Institutet KID Funding Program, the Swedish Research Council (grant. no. 621-2014-5503)
Human Frontier Science Program (grant. no. CDA-00033/2016-C)
Ragnar Söderberg Foundation (Fellows in Medicine 2016)
European Research Council under the European Union’s Horizon 2020 research and innovation programme (grant no. StG-2016_GENOMIS_715727)
Deciphering the role of programmed DNA breaks in aging
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