The total RNA is extracted from leaf and bulb of Lycoris radiata(L’Hér.)Herb.by an improved method of CsCl density gradient centrifugation.The suitable amount of initial sample is 1.0 g.In order to remove impuri...The total RNA is extracted from leaf and bulb of Lycoris radiata(L’Hér.)Herb.by an improved method of CsCl density gradient centrifugation.The suitable amount of initial sample is 1.0 g.In order to remove impurities and purify the total RNA,the crude extract of RNA dissolved in lysis buffer is extracted with the mixed solution of chloroform and isoamyl alcohol(V ∶V,24 ∶1),then the total RNA is precipitated.The purity,integrity and quality of the total RNA are detected by ultraviolet spectrophotometer,agarose gel electrophoresis and RT-PCR method,respectively.The results show that polysaccharide can be removed effectively from the total RNA extract by the improved method of CsCl density gradient centrifugation.The purity andintegrityof the total RNA of L.radiata are better,and the ratio of A260/A280 is 1.85-2.00.It is concluded that the total RNA of L.radiata bulb extracted by the improved method can satisfy the demand of molecular biology experiments.展开更多
本研究利用CTAB法提取感染枣疯病的枣树叶片总DNA,再通过氯化铯-双苯酰亚胺密度梯度离心从总DNA中富集纯化枣疯病植原体DNA。利用枣树26S r DNA基因和枣疯病植原体16S r DNA基因的特异引物,分别对富集纯化前后感病枣树叶片总DNA进行PCR...本研究利用CTAB法提取感染枣疯病的枣树叶片总DNA,再通过氯化铯-双苯酰亚胺密度梯度离心从总DNA中富集纯化枣疯病植原体DNA。利用枣树26S r DNA基因和枣疯病植原体16S r DNA基因的特异引物,分别对富集纯化前后感病枣树叶片总DNA进行PCR扩增。结果表明,模板浓度为1 ng/μL时,富集后的总DNA模板扩增枣树26S r DNA的电泳条带亮度低于富集前,扩增枣疯病植原体16S r DNA电泳条带亮度明显高于富集前。Real-time PCR检测结果表明,模板浓度为1 ng/μL时,纯化后的枣疯病植原体DNA中26S r DNA相对纯化前的量为0.0638,而JWB相对纯化前的量为0.5035,这说明采用氯化铯-双苯酰亚胺密度梯度离心法可以有效地从感染枣疯病枣树总DNA中富集纯化枣疯病植原体DNA。展开更多
文摘The total RNA is extracted from leaf and bulb of Lycoris radiata(L’Hér.)Herb.by an improved method of CsCl density gradient centrifugation.The suitable amount of initial sample is 1.0 g.In order to remove impurities and purify the total RNA,the crude extract of RNA dissolved in lysis buffer is extracted with the mixed solution of chloroform and isoamyl alcohol(V ∶V,24 ∶1),then the total RNA is precipitated.The purity,integrity and quality of the total RNA are detected by ultraviolet spectrophotometer,agarose gel electrophoresis and RT-PCR method,respectively.The results show that polysaccharide can be removed effectively from the total RNA extract by the improved method of CsCl density gradient centrifugation.The purity andintegrityof the total RNA of L.radiata are better,and the ratio of A260/A280 is 1.85-2.00.It is concluded that the total RNA of L.radiata bulb extracted by the improved method can satisfy the demand of molecular biology experiments.
文摘本研究利用CTAB法提取感染枣疯病的枣树叶片总DNA,再通过氯化铯-双苯酰亚胺密度梯度离心从总DNA中富集纯化枣疯病植原体DNA。利用枣树26S r DNA基因和枣疯病植原体16S r DNA基因的特异引物,分别对富集纯化前后感病枣树叶片总DNA进行PCR扩增。结果表明,模板浓度为1 ng/μL时,富集后的总DNA模板扩增枣树26S r DNA的电泳条带亮度低于富集前,扩增枣疯病植原体16S r DNA电泳条带亮度明显高于富集前。Real-time PCR检测结果表明,模板浓度为1 ng/μL时,纯化后的枣疯病植原体DNA中26S r DNA相对纯化前的量为0.0638,而JWB相对纯化前的量为0.5035,这说明采用氯化铯-双苯酰亚胺密度梯度离心法可以有效地从感染枣疯病枣树总DNA中富集纯化枣疯病植原体DNA。
