|Sarah Zhang working in the lab of Gerhard Bauer at UC Davis. She submitted this photo through Instagram to CIRM's #CIRMStemCellLab collection|
My name is Siruo Zhang and for the past eight weeks, I had been interning at the UC Davis Stem Cell Lab. As an intern, I worked under the instructions of Dr. Joseph Anderson and Sharlie Barclay from the HIV team in the lab.
My project is focused on testing for the safety of a combinatorial three gene anti-HIV vector. HIV, otherwise known as Human Immunodeficiency Virus, is a virus that targets and attacks T-cells and CD4 cells within the immune system. Over time, it begins to affect the body’s ability to ward off infections and can subsequently develop into AIDS. Statistics show that by the end of 2011, more than 34 million people worldwide were living with HIV.
Since all of the cells that HIV attacks come from hematopoietic stem cells, gene therapy becomes a reasonable option for treating and curing individuals of HIV. The combinatorial anti-HIV vector is a lentiviral vector that contains three components: the CCR5 shRNA, the TRIM5α, and the TAR decoy. Each component targets a specific stage of HIV’s life cycle, the CCR5 shRNA targets the pre-entry, the TRIM5α targets the post-entry/pre-integration, and the TAR decoy targets the post-integration. The CCR5 shRNA impedes the production of CCR5 by targeting the mRNA of CCR5, so that the viral envelope is unable to fuse with the cell membrane. The TRIM5α prevents virus uncoating, and the TAR decoy inhibits upregulation of HIV transcription.
In addition to these three anti-HIV genes, the vector also contain a CD25 pre-selective marker, which allows us to identify which cells contain the anti-HIV genes and which ones don’t. To test for the safety of this vector system, four experiments were conducted, QPCR testing for proto-oncogene expression, IL2 beta and gamma flow cytometry, CFU assay, and macrophage phenotype flow cytometry.
I enjoyed this internship immensely; I learned a lot of information about HIV and about stem cells. Before this internship, I hardly knew anything about stem cells, but now I can tick off the three properties that make a stem cell a stem cell. Those three properties are self-renewal, multi-potential, and highly proliferative. The most challenging part of this internship was the first week of being in the lab. I’ve never worked in an actual lab before, so everything was new to me. I messed up sometimes at first, but I learned from my mistakes and I tried to avoid it the next time. One of my favorite experiments that I conducted during my duration as an intern was probably plasmid extraction, because I’ve done it more times than I can count.
I would like to thank my mentors, Dr. Anderson and Sharlie Barclay, for being so patient with me and answering all of my questions. Also, I would like to thank Dr. Bauer for teaching me about stem cells. Lastly, I would like to thank CIRM and the UC Davis Stem Cell Program for providing me with this amazing opportunity. As my internship comes near its end, I can probably say that this summer was the most educational and enjoyable summer I’ve ever had. I gained not only knowledge, but also experience of working in a real lab.