浮游状态和生物膜状态白念珠菌总RNA提取方法的比较研究  

A comparative study on the extraction methods of Candida albicans in the form of planktonic and biofilm

作　　者：段薇 刘思琪 黄云 顾宁[1] 魏昕[1] DUAN Wei;LIU Siqi;HUANG Yun;GU Ning;WEI Xin(Department of Operative Dentistry and Endodontics,Afiliated Somatological Hospital of Nanjing Medical University,Jiangsu Province Key Laboratory of Oral Diseases,Jiangsuu Prouince Engineering Research Center of Stomatological Translational Medicine,Nanjing 210029,China)

机构地区：[1]南京医科大学附属口腔医院牙体牙髓科,江苏省口腔疾病研究重点实验室,江苏省口腔转化医学工程研究中心,江苏南京210029

出　　处：《口腔医学》2022年第5期416-419,430,共5页Stomatology

基　　金：国家自然科学基金(81970945)。

摘　　要：目的 本研究通过比较不同方法提取生物膜和浮游状态白念珠菌总RNA,以提供一种优化提取白念珠菌总RNA的有效方法。方法 将白念珠菌标准株SC5314在生物膜和浮游状态下培养24 h后,分别采用改良Trizol法和改良热酸酶酚法提取白念珠菌总RNA,用紫外分光光度计检测其纯度和浓度,琼脂糖凝胶电泳检测RNA的完整性,通过逆转录PCR反应验证总RNA的完整性及质量。结果 与改良热酸酶酚法相比,采用改良Trizol法提取白念珠菌总RNA操作时间较少(P<0.000 5),提取的总RNA纯度较高(P<0.05)。琼脂糖凝胶电泳显示两种方法提取的白念珠菌总RNA均呈现28S rRNA、18S rRNA和5S rRNA共3条清晰的条带,逆转录PCR反应也都获得清晰的目的条带。结论 改良Trizol法提取白念珠菌成熟期生物膜和浮游状态白念珠菌总RNA的纯度稳定,目前是一种高效、快捷、易于掌握的提取方法。Objective To provide an effective method to obtain total RNA of Candida albicans(C.albicans) in biofilm and planktonic states by comparing different extraction approaches. Methods Biofilms and planktonic C.albicans SC5314 were incubated for 24 h. Total RNA of C. albicans were extracted using modified Trizol method and modified hot phenol method, respectively. The difference in output, purity and integrity of total RNA extracted by two methods were compared by ultraviolet spectrophotometer, and further confirmed using agarose gel electrophoresis and RT-PCR. Results Modified Trizol method appeared more efficient than modified hot phenol method with less operation time(P<0.000 5). The purity of the total RNA extracted by modified Trizol method was higher than that by modified hot phenol method(P<0.05). The result of electrophoresis showed that there were three clear bands, 28 S rRNA, 18 S rRNA and 5 S rRNA in total RNA extracted by both modified Trizol method and modified hot phenol method. The result of RT-PCR confirmed the purity and integrity of total RNA of C.albicans with clear target bands. Conclusion Modified Trizol method contributes to more stable purity of the total RNA of C.albicans extracted in the form of planktonic than modified hot phenol method. Moreover, the modified Trizol method is rapid with easy operation and can be applied in experiments with efficiency.

关 键 词：白念珠菌 RNA提取 TRIZOL 生物膜 浮游 

分 类 号：R379.4[医药卫生—病原生物学]