槟榔叶表微生物前处理方式对基因组DNA提取效果的影响  

Effects of Genomic DNA Extraction on Microbial Pretreatment from Areca Leaf

作　　者：王贞霖 马瑞 徐芮 杨顺义 芮凯[3] Wang Zhenlin;Ma Rui;Xu Rui;Yang Shunyi;Rui Kai(Biocontrol Engineering Laboratory of Crop Diseases and Pests of Gansu Province,College of Plant Protection,Gansu Agricultural University,Lanzhou,730070;Research Center of the Quality Safety and Standards for Agricultural Products of Agricultural Sciences,Institute of Plant Protection,Hainan Academy of Agricultural Sciences,Haikou,571100;Sanya Institute,Hainan Academy of Agricultural Sciences,Sanya,572024)

机构地区：[1]甘肃农业大学植物保护学院,甘肃省农作物病虫害生物防治工程实验室,兰州730070 [2]海南省农业科学院植物保护研究所,海南省农业科学院农产品质量安全与标准研究中心,海口571100 [3]海南省农业科学院三亚研究院,三亚572024

出　　处：《分子植物育种》2024年第13期4341-4346,共6页Molecular Plant Breeding

基　　金：海南省自然科学基金青年基金项目(321QN365);海南省槟榔产业技术体系建设专项(HNARS)共同资助。

摘　　要：为了快速获取高质量的槟榔叶表微生物基因组DNA,本研究分别使用悬摇振荡和超声波洗脱微生物,使用真空抽滤和冷冻高速离心进行富集,利用超微量分光光度计对获取的叶表微生物DNA浓度和纯度进行测定,最后进行DNA电泳检测和PCR扩增检验对比提取效果。使用超声波法作为前处理时,所提基因组DNA浓度较低、纯度不在合格范围内;悬摇法提取的DNA浓度更高,且在悬摇20 min时所提基因组DNA浓度达最大值(25.67±0.96)μg/mL、(38.67±0.85)μg/mL;真空抽滤法较离心法而言,所提样本DNA浓度、纯度均更高。以悬摇振荡与真空抽滤相结合的方式为槟榔叶表微生物最佳收集方法,可以提高槟榔叶表微生物分离和收集的效率和质量,同时提高了叶表微生物DNA提取的成功率,本研究结果可为槟榔植物叶表微生物的收集提供参考依据。In order to rapidly obtain high-quality genomic DNA from areca leaf surfaces,this research utilized orbital shaking and ultrasonication to dislodge the microbes,followed by enrichment through vacuum filtration and high-speed centrifugation at low temperatures.The concentration and purity of the extracted microbial DNA were measured using a microvolume spectrophotometer.Finally,the effectiveness of the DNA extraction was compared through DNA electrophoresis and PCR amplification tests.When the ultrasonic method was used as a pretreatment,the concentration of genomic DNA was low and the purity was not within the acceptable range.The DNA concentration extracted by the orbital shaking method was higher,and reached its maximum values[(25.67±0.96)μg/mL and(38.67±0.85)μg/mL]after 20 minutes of shaking.Compared to the centrifugal method,the vacuum filtration method achieved higher DNA concentration and purity.The best method for collecting microorganisms from the surface of areca leaves was a combination of orbital shaking and vacuum filtration,which improved the efficiency and quality of microbial collection and also increased the success rate of DNA extraction from leaf surface microorganisms.The results of this study can provide a reference for the collection of microorganisms on the surface of areca leaves.

关 键 词：槟榔 叶表微生物 前处理 微生物收集 DNA提取 

分 类 号：S567.19[农业科学—中草药栽培]