摘要
目的探索白花蛇舌草总RNA最佳提取方法。方法以广东产白花蛇舌草为材料,选用Trizol法、多糖多酚总RNA提取试剂盒(RNA prep Pure Plant Kit)、植物总RNA提取试剂盒、改良CTAB法、RNAiso Plus法提取蛇舌草根和叶总RNA,经琼脂糖凝胶电泳及Aligent 2100检测,比较5种提取方法所得总RNA的完整性、纯度及产率。结果 RNA prep Pure Plant Kit及植物总RNA提取试剂盒均能提取到总RNA,1.5%琼脂糖凝胶电泳显示28S、18S条带清晰,前者亮度高于后者。RNA prep Pure Plant Kit中D(γ)_(260 nm)/D(γ)_(280 nm)和D(γ)_(260 nm)/D(γ)_(230 nm)值均在2.0左右,叶片组织RNA质量浓度为390.35 mg/L,经Aligent 2100检测,28S条带亮度是18S的2倍。改良CTAB法所得RNA 28S和18S条带稳定清晰,但受基因组污染严重。RNAiso Plus和Trizol法无法完成蛇舌草总RNA的提取。结论不同植物、不同品种或同一植物不同部位皆存在属性差异,要根据植物特性选择合适的RNA提取方法。RNA prep Pure Plant Kit提取蛇舌草根、叶片RNA效果较优,可满足后续试验要求。
Objective To study the best extraction method of total RNA in Hedyotis diffusa Willd.Methods Taking roots and leaves of Guangdong-grown Hedyotis diffusa Willd.as materials,Trizol method,RNA prep Pure Plant Kit,Plant Total RNA extraction kit,Improved CTAB method and RNAiso Plus Reagent method were used to extract total RNA,and the integrity,purity and yield of five total RNA extraction methods were compared by agarose gel electrophoresis（ AGE） and Aligent 2100.Results RNA prep Pure Plant Kit and Plant Total RNA extraction kit could extract the integral and high-quality total RNA.According the 1.5%agarose gel electrophoresis（ AGE） experiment,both the 28 S and 18 S bands were clear,and the former was brighter than the latter.D（γ）260 nm/D（γ）280nm and D（γ）260nm/ D（ γ）230nm were all in 2.0 above, and the leaves production rate was the highest and up to 390.35 g/L.The result of Aligent 2100 showed that the28 S band was twice the brightness as the 18 S.Although the improved CTAB method could effectively suppress the influence of polysaccharides and polyphenols, the genome were polluted seriously.Both RNAiso Plus reagent method and Trizol were useless for the extraction of total RNA.Conclusion The difference lies in different species of plants or different varieties of one species, and it even occurs in different parts of the same plant,so the RNA extraction method must be selected appropriately according to the characteristics of plants.RNA prep Pure Plant Kit has been proved to be able to extract good-quality RNA from the roots and leaves of Hedyotis diffusa Willd.tissue, which can meet the requirements of subsequent experiment.
出处
《广东药学院学报》
CAS
2016年第4期410-414,共5页
Academic Journal of Guangdong College of Pharmacy
基金
国家自然科学基金青年基金项目(81403195)
广东省自然科学基金项目(S2013010015418)
