摘要
目的:白木香次生代谢产物成分复杂,干扰其总RNA的提取,一般的提取方法无法获得总RNA或总RNA降解严重。为获得高质量的总RNA,本实验开展了白木香正常及结香组织采集及高质量总RNA提取方法的研究。方法:对从野外采集的白木香样品,用改良CTAB-LiCl法、去温浴的改良CTAB-LiCl法、SDS酸酚法、改良异硫氰酸胍-CTAB法和EASYspin RN09试剂盒5种方法分别对白木香叶、白木香枝条、白木香树干中的白木样品和结香样品进行RNA提取。结果:研究表明白木香枝条的离体样品室温保存5 h内不影响总RNA质量,确定EASYspin RN09试剂盒是白木香叶RNA的最佳提取方法,改良异硫氰酸胍-CTAB法是其他白木香组织RNA的最佳提取方法。结论:用改良异硫氰酸胍-CTAB法大量提取白木香树干中的白木样品及结香样品RNA,其28S∶18S为1.2～1.4,RIN值为6.8～7.5,并应用于转录组测序。
Objective：It is quite difficuh to isolate total RNA from Aquilaria sinensis due to the complexity of its secondary metabolites. In order to obtain the high-quality RNA, we investigated sampling method and RNA extraction from both normal and agilawood tissues of the plant. Methods ： Five methods such as modified CTAB-LiC1 method, modified CTAB-LiC1 method with no water- bath,SDS-acid phenol method, modified guanidinium thiocyanate-CTAB method and EASYspin RN09 RNA isolation kit were used to extract RNA from leaves, branches, white wood samples, agilawood samples of A. sinensis, respectively. Results：Total RNA was stable for 5 hours at room temperature after extracted from A. sinensis branches. EASYspin RN09 RNA isolation kit was the best extraction method for the leaf while modified guanidinium thiocyanate-CTAB method was the best for the other samples. Conclusion ： A great quantity of RNAs can be isolated from samples of white wood and agilawood wood, respectively. Their ratios of 28S： 18S are in the range of 1.2 - 1.4 and R1Ns are between 6. 8 - 7. 5. These RNAs can be applied in transcriptome sequencing.
出处
《中药材》
CAS
CSCD
北大核心
2013年第7期1055-1059,共5页
Journal of Chinese Medicinal Materials
基金
国家自然科学基金(31100496
81102418)
广东省中国科学院全面战略合作项目(2011B090300078)
广东省科技计划项目(2012A030100014)
关键词
白木香
总RNA提取
改良异硫氰酸胍-CTAB法
Aquilaria sinensis （ Lour. ） Gilg
Total RNA isolation
Modified guanidinium thiocyanate-CTAB method
作者简介
吴宏清（1987-），男，在读硕士研究生，专业方向：植物分子生物学；Tel：020．87688612，E—mail：whq0401@163．com。
通讯作者：白玲，Tel：0791-87529976，E-mail：bailing716@yall00．com．cn
章卫民，Tel：020-37656321，E—mail：wmzhallg58@qq．com。
