适用于厚朴的SRAP反应体系的建立与优化

利用正交试验L16(45),结合单因素试验对厚朴相关序列扩增多态性(Sequence-related amplified polymorphism,SRAP)标记反应体系的5个因素(Mg2+,dNTPs,引物,Taq酶和模板DNA)进行优化试验,结果表明:各因素水平变化对PCR反应的影响从大到小依次为:dNTPsTaq酶模板DNAMg2+引物;筛选出各反应因素的最佳水平,建立厚朴SRAP-PCR反应的最佳体系(25μL)为:Taq酶1.5 U,Mg2+1.8 mmol.L-1,模板DNA100 ng,dNTP 0.24 mmol.L-1,引物0.40μL。试验表明,该体系重复性好、稳定性强。

Establishment of SRAP-PCR Reaction Conditions in Magnolia officinalis

In this paper,the orthogonal design and single factor test design were used to optimize SRAP-PCR amplification system on Magnolia officinalis.in four levels of five factors(TaqDNA polymerase,Mg2+,DNA template,dNTPs,and primer,respectively),the results showed that the effects of each factors by the result of PCR were found.The order was dNTPs,TaqDNA polymerase,DNA template,Mg2+ and primer.A most suitable SRAP-PCR system for Magnolia officinalis was established,it was total 25 μL reaction system containing 1.5 U of TaqDNA polymerase,1.8 mmol·L-1 of Mg2+,100 ng of DNA template,0.24 mmol·L-1 of dNTPs,and 0.40 μmol·L-1 of primer.The new established SRAP-PCR system of Magnolia officinalis was with good repetition and stablility.This optimized system for SRAP marker would become one of the protocols for further research on Magnolia officinalis.