正交实验法优化千金藤属植物的DALP反应体系  

Optimizing DALP Reaction System of Stephania Yunnanenses Lo By Orthogonal Design

作　　者：马云淑[1] 李永谊[2] 虞泓[3] 

机构地区：[1]云南中医学院中药学院,云南昆明650200 [2]云南英茂生物技术实验室,云南昆明650106 [3]云南大学生命科学学院生态遗传学实验室,云南昆明650091

出　　处：《昆明医学院学报》2007年第4期31-34,共4页Journal of Kunming Medical College

摘　　要：目的建立一套稳定的千金藤植物DALP反应体系,为开展该属植物的DALP分子标记奠定基础.方法采用正交实验设计,以云南地不容Stephania yunnanensesLo与地不容Stephania epigaeaLo基因组DNA为模板,对DALP反应程序的重要参数进行优化.结果最佳的反应体系为20μL,包含模板DNA2.0μL,2.5 mmol/L的MgCl22.5μL,2.5 mmol/L的dNTPs 2.0μL,5 pmol/L的选择性引物1μL,5 pmol/L的反向引物3μL,0.5 U/μL的Taq酶5μL,10×Buffer 2.0μL,灭菌超纯水2.5μL.结论该DALP应用反应体系稳定、可靠、重现性好,可有效的地应用于千金藤植物的种间及居群间分类鉴定.Objective To establish a set of stable DALP reaction system of Stephania species and set foundation for DALP molecular markers of this genus. Methods Orthogonal design was used and the DNA of Stephania yunnanenses Lo and Stephania epigaea Lo were taken as samples to optimize some significant parameters of DALP reaction system. Results The 20 μL optimum DALP reaction system contains ： 2. 0 μL sample DNA, 2.5 mmol/L MgC122. 5 μL, 2.5 mmol/L dNTPs 2. 0 μL, 0. 5 U/μL Taq DNA polymerase 5 μL, 5 pmol/L selective primer 1 μL, 5 pmol/L reverse Primer 3 μL, 10 x Buffer 2 0 μL, and sterilized ultrapure water 2. 5 μL. Conclusion This reaction system is steady, reliable and repeatable. It is an effective reaction system for identifying species and local populations of Stephania species.

关 键 词：千金藤植物 直接扩增片段长度多态性 分子标记 反应体系 

分 类 号：R28[医药卫生—中药学]