The focus of the research in our laboratory is the transmembrane glycoprotein basigin. This interesting, multifunctional protein is involved in multiple cell processes, including embryonic development, embryo implantation into the uterus and metabolic regulation. Early studies demonstrated that basigin expression is elevated in rapidly growing cells such as cancer cells. This work suggested that releasing soluble basigin might stimulate the surrounding normal tissues to express a group of enzymes called extracellular matrix metalloproteinases (MMPs). MMPs play a central role in proliferation by breaking down the extracellular matrix surrounding layers of cells. We hypothesize that basigin expressed by cancer cells can induce MMP expression in normal tissues. To test this hypothesis, we sought to generate cell lines lacking a functional Basigin gene. Our lab used CRISPR/cas9 technology to attempt to disrupt the basigin gene (BSG) in T98 glioblastoma cells with the aim of eliminating basigin expression in these cells. For this, a Green Fluorescent Protein (GFP) DNA sequence was targeted for insertion into BSG. The work described herein shows the genetic analysis of the potential CRISPR/cas9 Basigin knockout clones using high-fidelity Polymerase Chain Reaction (PCR). Custom-designed oligonucleotide primers were used to amplify GFP DNA sequences from the T98 genomic DNA. The results of this study show that multiple independent T98 glioblastoma clones contain GFP DNA sequence in the genome. Future analysis of these clones will be performed to determine whether basigin protein expression was eliminated as a result of the CRISPR/cas9 targeting of the basigin gene.

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Faculty Advisor

Robert Belton

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