Generating GFP::HIS-35 Transgenic Strain through MosSCI Technique

The average human body contains around 50 to 75 trillion cells and each of these cells has about 1.6 meters of DNA that it needs to package inside of it. The cells are able to package the DNA using histones, which are basic proteins that keep DNA wound. Histones are also able to regulate genes important for development through the introduction of histone variants. In C. elegans, the canonical H2A histone has different variants such as HIS-35, which when mutated causes a decrease in fertility. This histone variant is only one amino acid different from H2A, which makes it very difficult to generate a HIS-35 specific antibody. To overcome this issue we have fluorescently tagged it with GFP. HIS-35 has been identified to be more enriched in sperm as opposed to embryos of C. elegans using mass spectroscopy. So my goal is to create this GFP::HIS-35 transgenic strain that will allow us to visualize where HIS-35 is localized in sperm. We have created a plasmid containing HIS-35 tagged with GFP in a bacterial vector which I will clone into another plasmid containing a working copy of a gene used as a selectable marker for introduction into C. elegans. I will then use a Mos 1 mediated Single Copy Gene Insertion (MosSCI) technique to create a single integrated copy of the GFP::HIS-35 transgene. Once we know where HIS-35 is localized then we can find at what time and place during sperm development it is found and how it affects fertility.