鲜榨果汁基因组DNA快速提取方法的建立及优化

Establishment and optimization of rapid genomic DNA extraction method for freshly squeezed fruit juice

中文关键词:  鲜榨果汁  滤纸片法  基因组 DNA  快速提取方法

英文关键词:freshly squeezed fruit juice  filter paper method  genomic DNA  rapid extraction method

中文摘要:

      目的 建立及优化鲜榨果汁基因组DNA快速提取方法。方法 本研究比较了植物DNA快速提取试剂盒法、棉签法、纤维素膜片法和滤纸片法4种方法提取15种鲜榨果汁基因组DNA的效果, 并筛选不同的提取液和裂解液、优化裂解液中的盐酸胍浓度和果汁基因组提取时间。结果 滤纸片法采用DNA裂解液(6 mol/L盐酸胍、1%聚乙烯吡咯烷酮、50 mmol/L三羟甲基氨基甲烷盐酸盐、20 mmol/L乙二胺四乙酸、21.3 mmol/L曲拉通-X-100; pH 6.4)、DNA漂洗液3 (8 mol/L盐酸胍、50 mmol/L三羟甲基氨基甲烷盐酸盐; pH 6.4)和DNA漂洗液4 (10 mmol/L 三羟甲基氨基甲烷盐酸盐、100 mmol/L氯化钠; pH 8.0)即可在5 min内从鲜榨果汁中提取得到适用于聚合酶链式反应(polymerase chain reaction, PCR)的基因组DNA。结论 本研究建立的滤纸片法不需要离心机等昂贵的仪器设备、操作简单、无刺激性气味、无环境条件限制, 具有快速、价廉、环保等优点, 为果汁基因组DNA快速提取与现场鉴定研究提供了参考。

英文摘要:

      Objective To establish and optimize a rapid method for extracting genomic DNA from freshly squeezed fruit juice. Methods This study compared the effectiveness of 4 kinds of methods: Plant DNA rapid extraction kit, cotton swab method, cellulose membrane method, and filter paper method for extracting genomic DNA from 15 types of freshly squeezed fruit juice. Different extracts and lysates were screened, the concentration of guanidine hydrochloride in lysates and the extraction time of fruit juice genome were optimized. Results The filter paper method, using DNA lysis solution (6 mol/L guanidine hydrochloride, 1% polyvinylpyrrolidone, 50 mmol/L trimethylol aminomethane hydrochloride, 20 mmol/L ethylenediamine tetraacetic acid, 21.3 mmol/L Tratone-X-100, pH 6.4), DNA bleach solution 3 (8 mol/L guanidine hydrochloride, 50 mmol/L trimethylol aminomethane hydrochloride, pH 6.4) and DNA bleach solution 4 (10 mmol/L trimethylol aminomethane hydrochloride, 100 mmol/L sodium chloride; pH 8.0), enabled the extraction of genomic DNA suitable for polymerase chain reaction (PCR) from freshly squeezed fruit juice within 5 minutes. Conclusion The established method does not require expensive equipment such as centrifuges, is simple to perform and free from any noxious odors, and is not constrained by environmental conditions. It offers advantages such as speed, affordability, and environmental friendliness, making it a valuable reference for rapid genomic DNA extraction and on-site identification research for fruit juices.