牛血清白蛋白对连香树PCR反应体系的优化  

Research on the Optimum PCR Reaction System of Cercidiphyllum japonicum with Bovine Serum Albumin

作　　者：黄绍辉[1] 方炎明[2] 

机构地区：[1]徐州工程学院,江苏徐州221008 [2]南京林业大学,江苏南京210037

出　　处：《安徽农业科学》2010年第26期14260-14261,共2页Journal of Anhui Agricultural Sciences

基　　金：江苏省农业三项工程项目[sx(2002)073]

摘　　要：[目的]为了取得更好的PCR扩增效果,建立高效的PCR反应体系。[方法]首先通过6因素5水平的正交试验对连香树RAPD-PCR反应体系优化组合,然后不再改变该结果中的其他条件,而仅改变Taq酶的浓度及在反应体系中加入不同浓度的牛血清白蛋白(BSA)。[结果]在25组Taq酶和BSA浓度组合中,BSA改善连香树RAPD-PCR反应的最佳浓度为0.6μg/μl,Taq酶浓度可由1.0μg/μl减少至0.4μg/μl。最后优化得到的20.0μl连香树RAPD反应体系为:25mmol/LMg2+2.0μl,dNTPs0.4μl,1U的TaqDNA酶1.0μl,10×Buffer缓冲液2.5μl,0.5OD引物0.8μl,约25ng模板0.6μl。[结论]BSA的适当加入减少了Taq酶的用量,在保证更好的试验效果前提下,节约了试验成本。[Objective] The efficient PCR reaction was established for its better result of PCR amplification.[Method] Firstly,the RAPD-PCR reaction system of Cercidiphyllum japonicum was optimized with the orthogonal design of 6 factors at 5 levels,and then,the concentration of Taq enzyme was only changed without any other variations in the experimental condition mentioned-above and the different concentrations of bovine serum albumin（BSA）were added for future experiment.[Results] In 25 combinations of Taq enzyme and BSA,the optimal concentration of BSA in the improvement of RAPD-PCR reaction of Cercidiphyllum japonicum was 0.6 μg/μl and the concentration of Taq enzyme could be decreased from 1.0 μg/μl to 0.4 μg/μl.Finally,the optimized RAPD reaction system for 20.0 μl Cercidiphyllum japonicum was：Mg2＋ of 25 mmol/L,2.0 μl;dNTPs,0.4 μl;Taq DNA enzyme of 1 U,1.0 μl;10 × Buffer buffer,2.5 μl;primer of 0.5OD,0.8 μl;and template of about 25 ng,0.6 μl.[Conclusion] The addition of BSA could reduce the appropriate amount of Taq enzyme for the better testing results with its cost-saving.

关 键 词：牛血清白蛋白 连香树 RAPD-PCR 反应体系优化 

分 类 号：S718.3[农业科学—林学]