罗红梅,苑天红,佘晓玲,贺娟,谭心,马研
LUO Hongmei,YUAN Tianhong,SHE Xiaoling,HE Juan,TAN Xin,MA Yan(Department of Microbiology

• 1:贵阳医学院微生物学教研室

摘要(Abstract)：

目的:探讨高效、简便提取白假丝酵母菌孢子相及菌丝相细胞总RNA的方法。方法:采用液氮法、超声法及玻璃珠(直径0.55 cm)法提取白假丝酵母菌的孢子相和菌丝相细胞总RNA,紫外分光光度计检测所提取的二相性白假丝酵母菌总RNA的OD260及OD280,琼脂糖凝胶电泳检测所提取的二相性白假丝酵母菌总RNA的完整性。结果:液氮法、超声法及玻璃珠法提取白假丝酵母菌孢子相总RNA的浓度分别为(1.021±0.145)g/L、(0.923±0.031)g/L、(1.121±0.175)g/L,纯度分别为1.697±0.087、1.474±0.064、1.897±0.047;液氮法、超声法及玻璃珠法提取白假丝酵母菌菌丝相总RNA的浓度分别为(0.357±0.260)g/L、(0.287±0.023)g/L、(0.523±0.056)g/L,纯度分别为1.548±0.047、1.474±0.064、1.874±0.064;电泳结果显示,应用玻璃珠法提取的孢子相和菌丝相总RNA 28S rRNA、18S rRNA、5S rRNA三条带均明显显示,而采用液氮法、超声法提取的总RNA三条带不明显。结论:RNAiso Plus试剂联合直径为0.55 cm的玻璃珠法提取的白假丝酵母菌孢子相及菌丝相细胞的总RNA浓度及纯度均较液氮法和超声法好,值得推广。
Objective: To explore an efficient and simple method for extraction of total RNA of Candida albicans in yeast form and hyphal form.Methods: The total RNA of Candida albicans in yeast form and hyphal form was isolated by liquid nitrogen method,ultrasonication method and glass bead(diameter 0.55 cm)method respectively.The OD260 and OD280 values of extracted total RNA were measured by ultraviolet spectrophotometer and agarose gel electrophoresis was carried out to test RNA integrity.Results: The concentrations of yeast form total RNA extracted by liquid nitrogen method,ultrasonication method and glass bead method were(1.021±0.145)g/L,(0.923±0.031)g/L,(1.121±0.175)g/L,and the purities were 1.697±0.087,1.474±0.064,1.897±0.047,respectively.The concentrations of hyphal form total RNA extracted by the three methods were(0.357±0.260)g/L,(0.287±0.023)g/L,(0.523±0.056)g/L,and the purities were 1.548±0.047,1.474±0.064,1.874±0.064,respectively.The results of electrophoresis showed that there were three clear bands-28S rRNA,18S rRNA and 5S rRNA when total RNA was isolated with glass bead method and the bands were not clear when RNA was isolated by liquid nitrogen and ultrasonication methods.Conclusions: The concentration and the purity of total RNA isolated from dimorphic Candida albicans by RNAiso Plus combined with 0.55 cm diameter glass bead method are higher than that isolated by liquid nitrogen and ultrasonication methods.This method is effective,simple,economic and worth popularizing.

关键词(KeyWords)： 白假丝酵母菌;RNA;液氮;超声;玻璃珠
Candida albicans;RNA;liquid nitrogen;ultrasonication method;glass beads

Abstract:

Keywords:

基金项目(Foundation): 贵州省科学技术厅科研基金资助项目【2008】2273号

作者(Author): 罗红梅,苑天红,佘晓玲,贺娟,谭心,马研
LUO Hongmei,YUAN Tianhong,SHE Xiaoling,HE Juan,TAN Xin,MA Yan(Department of Microbiology

DOI: 10.19367/j.cnki.1000-2707.2012.06.009

参考文献(References)：

• [1]Tashiro M,Murakami H,Yoshizawa S,et al.Isolationrate and susceptibilities of Candida species from blood,vascular catheter,urineand stool[J].KansenshogakuZasshi,2010(2):187-192.
• [2]苏惠春,程波,傅冷西,等.3种破壁方法提取念珠菌总RNA效果的比较[J].中国真菌学杂志,2007(5):286-288.
• [3]潘琤,魏昕,刘卫红.改进Trizol法提取白假丝酵母菌总RNA初探[J].临床口腔医学杂志,2007(9):531-532.
• [4]苏惠春,程波,施秀明.EFG1和HGC1基因在白假丝酵母菌菌丝相和酵母相的表达差异[J].中国皮肤性病学杂志,2010(4):304-306.
• [5]韩慧霞,王鲁,宋秋荷,等.白假丝酵母菌菌丝相全长cDNA文库的构建和鉴定[J].中国皮肤性病学杂志,2011(4):260-263.
• [6]陈雪蓉,肖敦振.白假丝酵母菌毒力因子的研究进展[J].中国妇幼保健,2007(32):4631-4633.
• [7]Chaffin WL,Lopez-Ribot JL,Casanova M,et al.Cellwall and secreted proteins of Candida albicans:Identifi-cation,function and expression[J].Microbiol Mol BiolRev,2008(1):130-180.

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