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Trizol试剂法快速高效提取3种作物不同组织总RNA
作者: 吴凯朝1  2  3  黄诚梅3  李杨瑞1  2  3 *  杨丽涛1  2 *  吴建明2  
单位: (1 广西大学/广西亚热带生物资源保护和利用重点实验室   南宁 530005;2 中国农业科学院甘蔗研究中心/广西农业科学院甘蔗研究所/  
关键词: RNA  Trizol试剂法  改良Trizol-异丙醇法  甘蔗  水稻  玉米  
分类号:Q781
出版年,卷(期):页码:2012 ,43 ( 12 ): 页码:1934-1939
摘要:

【Objective】The present study was conducted to establish a fast and effectively method for isolating the total RNA in gramineous crops in order to provide foundation for subsequent molecular biology researches. 【Method】Using the Trizol reagent method, the supernatant amount changed. Then, by the precipitation method and precipitation rinse times, the total RNA extraction from the leaves, stem tips, and roots of sugarcane, maize, and rice was optimized. 【Result】Compared with the 0.4 mL supernatant absorbent, the 0.2 mL supernatant absorbent’s total RNA OD260/OD280 and OD260/OD230 values were between 1.95-2.00 and 2.11-2.44 respectively, in which the total RNA obtained was highly purified. The total RNA OD260/OD230 value (2.11-2.47) that underwent precipitation rinse two times was significantly higher than the total RNA OD260/OD230 value (1.01-1.61) that underwent precipitation rinse 1 time, so the total RNA of the higher OD260/OD230 value was purer. The spectra of total RNA 28S, 18S, and 5S resulted from the isopropanol precipitation in various materials were clear and without trailing; therefore the total RNA integrality was better. However, from the LiCl precipitation method, the resulting RNA band was adhesively fuzzy and the 5S band was apparently diffused, so the total yield of RNA was relatively low. Utilizing the Trizol-isopropanol extraction method, the total RNA from sugarcane was extracted as template for GAPDH gene fragmentation, and the resulting GAPDH gene expressions obtained from sugarcane leaves, stalk tips, and root tips were very strong with no specific amplifications. Hence, the objective-to-fragment length was 153 bp. 【Conclusion】The total RNA tissues extracted from sugarcane, maize, and rice through the modified Trizol-isopropanol extraction method showed clear bands, good integrity, high purity, and high yield. Therefore, this extraction method is suitable for the fast and efficient extraction of total RNA from different tissues of gramineous plants.

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