Ethanolfixatie en DAPI-kleuring: een methode om DNA in C. elegans te visualiseren

- Start with a drop of buffer solution on a microscope slide and transfer intact worms to the droplet. Once the worms are in place, use a lint-free tissue to soak up and remove the buffer. Fix the sample by bathing the specimens with 90% ethanol several times, allowing it to evaporate after each application.

Once the ethanol has evaporated after the final rinse, add a solution containing the nucleic acid dye, DAPI, to the sample. DAPI binds preferentially to adenine and thymine bases in the minor groove of DNA. When bound, the DAPI-DNA complex absorbs UV light and emits visible blue light.

Next add an anti-fade preservative to extend the shelf life of the sample. Apply a cover slip and seal it to the slide. Then use a fluorescent microscope with a blue filter to visualize DAPI bodies, which, depending on the cell and its cycle status, can represent single chromosomes, sister chromatid pairs, or whole nuclei. In this experiment, we will count DAPI stained sister chromatid pairs in oocytes, to identify tetraploid strains of Caenorhabditis elegans.

- Tetraploid strains have 12 chromosome pairs, which can be validated by counting the DAPI stained bodies in unfertilized oocytes. To do s