Katie Munechika / Dr. Ethel Cesarman - Week 1

During this first week, I have been focused on becoming familiar with the new lab environment and learning the protocols for the experiments I will need to perform. The project I am working on is studying the involvement of linker histone H1 mutations on lymphoma development. In particular, we are focused on H1 isoforms H1C and H1E, as those are the most commonly mutated isoforms in human lymphoma. Current antibodies for these isoforms are not very effective, so in order to perform an immunoprecipitation of these, they require a protein tag with reliable antibodies to be added to the N or C terminal of the target protein. Recently, a post-doc in the lab has successfully added a V5 tag to the N-terminal of H1E isoform in lymphoma cells (confirmed by sequencing). The main task I have been working on this week is verifying that the tag is able to be used to pull-down H1E. I did this by testing the V5 antibody with the tagged H1E cell lysates via a western blot. Unfortunately, the western blot was unsuccessful, as the V5-tagged H1E band did not appear. I believe the reason for this may have been that the V5 antibody used was too old, so I am planning to try again with a new antibody.

Additionally, I have been helping with some mouse work, which is a new experience for me. We currently have mice that have been genetically altered to contain H1C and H1E mutations, so I worked on extracting genomic DNA from mouse tails for genotyping. I also helped with bone marrow extraction from some of the mice to be used for bone-marrow transplants in later experiments focused on the effects of H1C and H1E mutations on lymphoma development.

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