摘要
RNA试剂盒法是获取植物高质量RNA样品的常用技术,针对3种常见RNA提取试剂盒在红地球葡萄叶样品RNA提取效果进行评估和优化,以期找到适合红地球葡萄叶片RNA表达谱测序需求的高质量RNA提取策略。试验依据3种试剂盒推荐时间和条件进行RNA提取,并对时间等条件进行优化,比对了优化前后RNA质量。结果显示,试剂盒经优化后的方法提取的RNA收率高,完整性好,OD260/OD280在1.8-2.2之间,OD260/OD230大于2,28S与18S条带清晰完整,RNA-Seq测序质量及基因覆盖度符合要求。在红地球葡萄叶片上的RNA提取结果表明,延长提取试剂与样品的接触时间操作可以显著提高RNA提取质量(纯度/完整性)。
Commercial RNA extraction kits were wildly used in obtaining high quality RNA samples from plants.To seek the optimal method for the extraction of high quality RNA in RNA Expression Sequences Array（RNA-Seq）,the topic of present article is to evaluate and optimize the effects of three common RNA extraction kits on leaves samples from grapevine（Vitis vinifera cv.Red Globe）.The quantities of RNA were analyzed between the recommendatory and optimal usages of three kits in term of times.Results showed that the optimized procedure for RNA extraction displayed higher yield and better integrity on total RNA.There are a ratios of OD260/OD280 from 1.8 to 2.2 and further OD260/ OD230ratio no less than 2,resulted in a integrate 28S and 18S bands by electrophoresis.The extracted samples also kept their quality and gene coverage from RNA-Seq sequencing.Our research demonstrated that the prolonged incubation time for samples preparation and other procedures would significantly benefit the separation of total RNA from grape leaves（Vitis viniferacv.Red Globe）with a higher purity and integrity.
出处
《生物技术通报》
CAS
CSCD
北大核心
2013年第6期215-220,共6页
Biotechnology Bulletin
基金
国家自然科学基金项目(30860032
31160058)
兵团博士资金项目(2011BB018)
