The construction and functional study of ORF19.2730 recombinant DNA of Candida albicans

• Main Authors: Sheng-Shyan Tai, 戴勝賢
• Other Authors: Yun-Liang Yang
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/2pqt2u

碩士 === 國立交通大學 === 理學院碩士在職專班網路學習學程 === 96 === Candida albicans is one of the important opportunistic fungal pathogens in human body. It has various morphologies. When body immunity deteriorates or microbial ecology in the body changes due to medical reasons, the fungus would have opportunity to infect mucous tissue; typical clinical symptoms include thrush and vaginitis. In the worst case, it would lead to general system infection and death. Recently, because of the increase of immune deficiency patients and drug resistance due to improper use of antibiotics, the cases of nosocomial infection keeps rising. Genetic and functional studies on Candida albicans may help in issues of infection prevention, treatment, and drug resistance. Previously, the laboratory has conducted suppression subtractive hybridization and found the sequence of orf19.2730 might have involved in the morphogenesis of Candida albicans. Orf19.2730 is a hypothetical gene consisting 1821 nucleotides and is in Contig19-10158 on the fourth chromosome of C. albicans. The homologous allele is orf19.10244. The sequence of orf19.2730 and the upstream promoter, 2192 nucleotides in total, was amplified by PCR. The ORF was then cloned into 2μ vector for expression in Saccharomyces cerevisiae. The promoter sequence was fused to lacZ gene and then was transformed into Saccharomyces cerevisiae for the purpose of identifying cis-acting sequences regulating the promoter activity. Western blot analysis showed that the ORF indeed could manifest, able to produce protein. β-galactosidase filter assay also showed that there may have inhibitory sequence from -257 to -315 nucleotides.