CAJ | 학술논문

目的当经典Trizol法提取骨组织RNA不能满足实验要求时,使用盐溶液再处理提取的一种改良方法.方法对经典Trizol法提取的骨组织RNA,用异丙醇再沉淀时,加入柠檬酸钠、氯化钠混合盐溶液溶解,再抽提RNA.测定RNA纯度、浓度和完整性,聚合酶链反应(PCR)鉴定其有效性.结果重新提取的RNA能够被焦磷酸二乙酯(DEPC)水充分溶解,盐溶液处理后的RNA浓度与处理前相近;处理后RNA的A260/A280比值、A260/A230比值分别为1.96 ±0.11、1.90 ±0.21,均分别显著高于处理前(1.52±0.05、0.27 ±0.08);凝胶电泳示RNA完整性,逆转录后可扩增出目的基因.结论改良Trizol法重提的总RNA纯度较常规法显著提高,且完整性好,可以满足进一步分子研究的要求.
목적 당경전Trizol법제취골조직RNA불능만족실험요구시,사용염용액재처리제취적일충개량방법.방법 대경전Trizol법제취적골조직RNA,용이병순재침정시,가입저몽산납、록화납혼합염용액용해,재추제RNA.측정RNA순도、농도화완정성,취합매련반응(PCR)감정기유효성.결과 중신제취적RNA능구피초린산이을지(DEPC)수충분용해,염용액처리후적RNA농도여처리전상근;처리후RNA적A260/A280비치、A260/A230비치분별위1.96 ±0.11、1.90 ±0.21,균분별현저고우처리전(1.52±0.05、0.27 ±0.08);응효전영시RNA완정성,역전록후가확증출목적기인.결론 개량Trizol법중제적총RNA순도교상규법현저제고,차완정성호,가이만족진일보분자연구적요구.
Objective To introduce one method to improve the effect of bone RNA extraction when the classical Trizol method can not meet the experimental requirements.Methods The concentration,purity and integrity of the extracted RNA were tested.The RNA was reversely transcripted into cDNA,as a template which was then amplified to DNA.Results The re-extracted RNA could be fully dissolved by diethyl chlorophosphate (DEPC) water.This method could effectively reduce the interference of mucin carbohydrate.After the prevention of the sodium mixture solution,the concentration of the RNA was slightly lower than that before the prevention.The rate of A260/A280 and A260/A230 of RNA was both significantly higher afher the prevention than that before the prevention [(1.96 ± 0.11) vs.(1.52 ± 0.05),(1.90 ±0.21) vs.(0.27 ±0.08) respectively].Agarose gel electrophoresis showed the RNA after prevention was complete and the polymerase chain reaction (PCR) procession could successfully amplify the targeted gene.Conclusion The improved extraction method could insure the purity,concentration and integrity of RNA,and meet the requirements for molecular research.