建構熱帶念珠菌DNA表達文庫

Abstract

念珠菌（Candida spp.）是伺機性病原真菌，在免疫低下的病患身上會造成念珠菌症。念珠菌中以白色念珠菌所造成的感染是最多的。值得一提的是，熱帶念珠菌( Candida tropicalis )是台灣最普遍的非白色念珠菌致病性真菌。雖然有抗真菌藥劑可以處治念珠菌感染，但近年來抗藥菌株的出現，卻是個日漸嚴重的問題。其中如最主力使用的fluconazole 也出現抗藥菌株。fluconazole 抗藥性的存在，使其對治療造成嚴重的威脅。為了之後的功能性研究，例如發病機理和抗藥性，我利用pYES3/CT這個酵母菌表現載體，將熱帶念珠菌的 genomic DNA clone 進去，並確認此文庫的效率及複製的能力，建構出兩個基因組DNA 表達文庫和一個局部性DNA 表達文庫。之後並利用這些文庫進行 miconazole 抗性基因及侵犯力基因的篩選，得到可能與 miconazole 抗性有關的基因。這些基因的功能有 DNA 複製起始和 citrate synthase。未來，研究人員可以藉由分析這個文庫，來找出其他參與抗藥性和發病機制的新基因，並可成為潛在的藥物靶點。

Candida species are opportunistic yeast pathogen that can cause candidiasis in immunocompromised patients. Among them, Candida albicans is the most dorminant ones. It is worth mentioning that Candida tropicalis is the most prevalent pathogenic yeast species of the non-albicans Candida in Taiwan. The emergence of fluconazole-resistant C. tropicalis makes this pathogen a potential clinical threat. To further functional studies, such as pathogenesis and drug resistance, I used pYES3/CT, a S. cerevisiae expression vector, to clone the genomic DNA of C. tropicalis into an expression library. Finally, I constructed two expressing genomic DNA libraries and a partial one. After screened these libraries for miconazole resistance and invasive ability genes, I found several candidats, two of them are associated with miconazole resistance. Those genes are suitable for studying drug resistance and pathogenesis.