外周血基因组DNA提取试剂盒的改良  

Improvement of Peripheral Blood Genomic DNA Extraction Kit

作　　者：陶莹 廖长秀 覃宇馨 何南轩 TAO Ying;LIAO Zhang-xiu;QIN Yu-xin;HE Nan-xuan(School of Pharmacy,Youjiang Medical College for Nationalities,Guangxi Baise 533000;School of Basic Medicine,Youjiang Medical College for Nationalities,Guangxi Baise 533000;School of Medical Imageology,Youjiang Medical College for Nationalities,Guangxi Baise 533000,China)

机构地区：[1]右江民医学院药学院,广西百色533000 [2]右江民族医学院基础医学院,广西百色533000 [3]右江民族医学院医学影像学院,广西百色533000

出　　处：《广州化工》2024年第2期140-142,共3页GuangZhou Chemical Industry

基　　金：国家自然科学基金(81660642);2022年度广西学位与研究生教育改革课题(JGY2022278)。

摘　　要：对现有DNA提取试剂盒进行改良,减少DNA提取成本,寻找经济、稳定的提取方法。将96份血液标本分为A、B两组,各48份,组内按外周血体积分为250μL、500μL、750μL三组。A组(试剂盒法)采用杭州倍沃医学科技公司的血液基因组小量提取试剂盒(离心柱型),B组(改良法)使用3%明胶吸附白细胞代替试剂盒法中分离白细胞的步骤,其余步骤同试剂盒法。采用紫外分光光度计、凝胶电泳、荧光定量PCR比较两种提取方法不同体积外周血DNA提取效果。与试剂盒法相比,改良法250μL DNA提取得率稍低,差异无统计学意义(P>0.05);改良法500μL、750μL DNA提取得率及500μL的OD_(260)/OD_(280)值均显著高于试剂盒法(P<0.01),差异具有统计学意义。琼脂糖凝胶电泳发现两种方法提取的DNA条带均完整,无拖尾现象,定量PCR扩增后两者产物融解曲线均为单峰,Ct值差异无统计学意义。本实验中改良全血DNA提取法在提取大量血体积DNA时具有优势,是一种经济、稳定、高效的DNA提取方法。To reduce the cost of extraction and find an economic and stable method of peripheral blood DNA extraction,the method of DNA extraction kit was improved.96 blood samples were divided into two groups with 48 samples in each group,and then each group was divided into 250μL,500μL and 750μL three blood volume.Peripheral blood DNA was extracted by Biomiga Blood gDNA Isolation Kit(Column)in kit method.3% gelatin was used toadsorb leukocytes in improved method instead of the steps of separating leukocytes in the kit extraction method.The quality of DNA extraction from peripheral blood with different volumes by the two extraction methods was observed by UV spectrophotometer,gel electrophoresis and fluorescence quantitative PCR.The yields of DNA extracted in improved method with 250μL peripheral blood were slightly lower than those in kit method with no statistical significance(P>0.05).The yields of DNA extracted in improved method with 500μL and 750μL peripheral blood were significantly higher than those in kit method(P<0.05).Moreover,the value of OD_(260)/OD_(280) in improved method with 500μL peripheral blood was higher than that in kit method(P<0.01).Using agarose gel electrophoresis,there was a single and complete band without tailing for DNA extracted from different volumes of peripheral blood by two methods.In addition,the melting curve peak was single,and there was no difference in Ct value for fluorescent quantitative PCR of extracted DNA from different volumes of peripheral blood by two methods.The improved method of peripheral blood DNA extraction had advantages in extracting large amounts of blood volume DNA,it was a stable,economical and efficient method for extracting large blood volumes.

关 键 词：外周血 基因组DNA 提取 改良 

分 类 号：R34[医药卫生—基础医学]