Open Access Honors Program Thesis
James Jurgenson, Honors Thesis Advisor
Fusarium--Genetic engineering; CRISPR-associated protein 9;
In molecular biology, CRISPR-Cas9 is a useful gene editing tool that takes advantage of a powerful cleaving enzyme and DNA sequences found in prokaryotes. In this study, a target-specific CRISPR-Cas9 vector containing an RNA-guided Cas9 nuclease specific for the FUM1 gene was constructed for the purpose of introducing mutations into the FUM1 gene of Fusarium verticillioides. Plasmids pFC332 and pFC334 were isolated. A guide RNA sequence was designed and amplified using Polymerase Chain Reaction (PCR) with pFC334 as the template. Sticky ends were created through restriction enzyme digestion of pFC332. These pieces were combined using Uracil-Specific Excision Reagent (USER) fusion, and the resulting vector was transformed into competent E. coli cells. Colonies containing the constructed vector were selected, and PCR was performed for amplification. The results were purified and stored at - 40℃. This constructed vector can be used for the specific mutation of the FUM1 gene of Fusarium verticillioides.
Year of Submission
Department of Biology
University Honors Designation
A thesis submitted in partial fulfillment of the requirements for the designation University Honors
1 PDF file (21 pages)
©2020 Jennifer Petsche
Petsche, Jennifer, "Construction of a target-specific CRISPR-Cas9 vector for the purpose of introducing mutations into the FUM1 gene of Fusarium verticillioides" (2020). Honors Program Theses. 442.