关键词: 新生隐球菌, RNA, 真菌, 液氮, Yeast RNA kit, 酸洗玻璃珠法

Abstract: Objective A quick approach to extract high quality total RNA from Cryptococcus neoformans plays a vital role in researches on the molecular detection and disease-related gene expression of high pathogenic Cryptococcus neoformans.Method Cryptoccocus neoformans strain and Cryptoccocus neoformans capsule deficient strain were used in the research. Four methods were designed to perform the extraction: acid washed glass beads approach, liquid nitrogen approach, guanidinium isothiocyanate one step approach and acid washed glass beads associated with Yeast RNA kit approach. RNA quality is determined by measurement of OD260 and OD280 with UV spectrophotometer, by agarose gel electrophoresis and by quantitative PCR.Result RNA yield of the four approaches, namely, acid washed glass beads approach, liquid nitrogen approach, guanidinium isothiocyanate one step approach and acid washed glass beads associated with Yeast RNA kit approach, was 0.2 μg/10⁵ cells, 0.4 μg/10⁵ cells, 0.1 μg/10⁵ cells and 0.6 μg/10⁵ cells respectively.Conclusion RNA which gained by the approach of acid washed glass beads associated with Yeast RNA kit has the best homogeneity and integrity. Thus, this method is ideal to perform the extraction of total RNA from Cryptococcus neoformans under protection of capsule and cell wall.

Key words: Cryptoccocus neoformans, RNA, fungus, liquid nitrogen, Yeast RNA kit, acid washed glass beads approach