Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of autoantibodies against self-antigens. SLE has a complex multifactorial genetic basis. Genome wide association studies (GWAS) have implicated polymorphisms causing decreased expression in many genes in SLE etiology and will likely continue to implicate new genes. Therefore, the aim of my project was to create a modular system where the effect of knocking down a gene of interest can be studied in cell culture in a timely manner. This was accomplished by inserting shRNAmirs targeting our validation gene, A20, into a retroviral vector, creating viral particles using a packaging cell line, and infecting cultured cells. Two shRNAmirs were determined to be effective by qRT-PCR and Western blots on infected cells. Vectors carrying these shRNAmirs were used to infect ex vivo B cells and BMDCs, and results suggested that A20 knockdown may mediate functional changes in both cell types.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/43083 |
| Date | 04 December 2013 |
| Creators | Lifeso, Kimberley |
| Contributors | Wither, Joan |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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