提取航天诱变向日葵种子总RNA的一种有效方法

A Method for Efficient Extraction of Total RNA from Seeds of Sunflower Induced by Space Flight

目的:从航天诱变向日葵种子中提取高质量的总RNA。方法:采用改进的SDS法,提取缓冲液与氯仿同时作用液氮研磨材料后,用酸酚-氯仿抽提一次,经LiCl过夜沉淀、DNaseⅠ处理、1/2体积的无水乙醇沉淀多糖,最后加入1/10体积的醋酸钠和2倍体积的无水乙醇沉淀总RNA,用琼脂糖凝胶电泳与紫外分光光度法测定产量与纯度,用PT-PCR鉴定RNA的质量。结果:提取的RNA电泳条带清晰,D260nm/D230nm、D260nm/D280nm值分别为2.09、2.00,其产量为35.30μg/100 mg,用PT-PCR能扩增出肌动蛋白基因片段。结论:采用该方法提取的总RNA纯度高、完整性好,适于进一步的分子生物学研究。

Objective：To obtain total RNA with high quality from sunflower seeds induced by space flight.Methods：The grind powders in liquid nitrogen were transferred into extraction buffer containing SDS with equal volume of chloroform for extraction,and then extracted by acidic phenol,followed by precipitation overnight by LiCl.After digested by DNaseⅠ,polysaccharide was removed by precipitation with 1/2 volume of absolute ethanol,and finally the total RNA was precipitated with 1/10 volume of NaAc and 2 volume of absolute ethanol.The yield and purity of the total RNA was tested by electrophoresis and spectrophotometry,and its quality was identified by RT-PCR.Results：The electrophoretic bands of the extracted RNA were clear.The D260nm/D230nm and D260nm/D280nm ratio for total RNA were 2.09 and 2.00 respectively.And the yield of RNA was 35.30 μg/100 mg.Fragment of actin gene could be amplified by RT-PCR from the extracted total RNA.Conclusion：Total RNA extracted by the modified method was of high purity and integrity,and could be used for further molecular s