A Heterologous Screen of the Neisseria gonorrhoeae Genetic Library and its Effects on the Expression of . the Gene of Unknown Function, NG 1684
Wood, Brianne M.
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Neisseria gonorrhoeae is a well-known pathogen in today's society. The bacterium is the cause of the sexually transmitted disease gonorrhea, and if left untreated can lead to serious sequellae. Its success lies in its pathogenicity; additionally it is antibiotic resistant. Although the disease has been around for thousands of years there is still a lot unknown about the virulence of N gonorrhoeae. One particular study used a tissue model culture for gonoccocal infection to show that one individual gene's expression is the most up regulated once N gonorrhoeae has adhered to host epithelial cells; that gene is NG 1684. The purpose of our study was to examine this gene more closely. Its expression is linked to adhesion, but the exact function of the gene is unknown. In order to find an explanation for this a heterologous genetic screen was performed using Escherichia coli strain P90C modified with a NG 1684-lacZ fusion insert, and then screened with the N gonorrhoeae pCBB partial DNA plasmid library. Expression of NG 1684 was studied qualitatively by growing the bacteria on Xgal plates where colonies would turn varying shades of blue due to the presence of the reporter gene lacZ and its activity. Expression was also measured quantitatively by using the reporting effects of lacZ gene with β -galactosidase activity assays. Isolates producing different activities than the empty vector negative control were purified and sequenced. A total of five plasmid inserts were sequenced and each encoded different inserts of the N. gonorrhoeae genome. Initially plasmid pBW03 was the most intriguing as it partially encoded for a regulator. More studies are necessary, such as developing knock outs to make mutant strains of N. gonorrhoeae in order to determine the effects on NG 1684 expression in its natural context.