Acta Agrestia Sinica

›› 2006, Vol. 14 ›› Issue (4): 379-383.DOI: 10.11733/j.issn.1007-0435.2006.04.017

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Optimization of SCAR Reaction System for Alfalfa

REN Wei-bo1, HAN Jian-guo1, GUO Hui-qin2, FANG Cheng1, YANG Qin-chuan3   

  1. 1. Institute of Grassland Science, China Agriculture University, Beijing 10094, China;
    2. Institute of Bioengineering Science, Inner Mongolia Agriculture University, Huhhot, Inner MongoliaAutonomous Region 010018, China;
    3. Institute of Animal Science, CAAS, Beijing 100094, China
  • Received:2006-04-19 Revised:2006-11-17 Online:2006-11-15 Published:2006-11-15

紫花苜蓿耐盐测序扩增区段标记条件优化

任卫波1, 韩建国1, 郭慧琴2, 方程1, 杨青川3   

  1. 1. 中国农业大学草地研究所, 北京, 100094;
    2. 内蒙古农业大学生物工程学院, 呼和浩特, 00000;
    3. 中国农业科学院北京畜牧兽医研究所, 北京, 100094
  • 通讯作者: 韩建国,E-mail:grasslab@public3.bt a.n et.cn
  • 作者简介:任卫波(1979- ),男,山西晋城人,博士研究生,主要研究方向为牧草育种
  • 基金资助:
    北京市教委共建项目(XK100190552;JD100190531)

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Abstract: Alfalfa cultivar “Zhongmu No.1” was used as materials for extracting DNA in present experiment. The gradient tests on the concentration of Mg2+, dNTP, SCAR primer, template DNA, TaqDNA polymerase were conducted. Results show that two of three primer sets could amplify target bands, among which SCAR1 was the suitable primer pair. The stable reaction system of SCAR with a total volume of 25 μl, consisting of Mg2+ of 2.5 mM concentration, dNTP at 200 μM, SCAR primer at 0.5 μM, TaqDNA polymerase at 1.5 U, and 50～100 ng template DNA. A clear, stable band can be amplified with the optimized reaction system which can be used in molecular marker diagnoses of salt tolerance of the alfalfa populations.

Key words: Alfalfa, SCAR, PCR amplified reaction optimization

摘要： 以中苜一号紫花苜蓿(Medicago sativa L.cv.Zhang mu No.1)为材料,对SCAR反应中的引物浓度、退火温度、Mg2+、dNTP浓度、模板DNA用量及Taq酶用量进行优化。结果表明:在3对SCAR中有2对引物出现目标扩增带,其中以SCAR1的效果最好;适宜反应体系总体积为25μL、Mg2+浓度2.5mM、dNTP浓度200μM、引物浓度0.5μM、Taq酶1.5U、模板DNA为50~100ng;优化后的反应体系能得到清晰稳定的SCAR扩增条带,可用于苜蓿耐盐性分子标记鉴定分析。

关键词: 紫花苜蓿, 耐盐SCAR, PCR条件优化

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