WANG Xindi,HUANG Rukui,FENG Chengcheng,et al.Orthogonal Design-direct Analysis of SSR-PCR Reaction System in Momordica charantia[J].Northern Horticulture,2016,40(10):99-103.[doi:10.11937/bfyy.201610025]
正交设计直观分析法优化苦瓜SSR-PCR反应体系
- Title:
- Orthogonal Design-direct Analysis of SSR-PCR Reaction System in Momordica charantia
- 文献标志码:
- A
- 摘要:
- 以苦瓜‘MC9’为试材，采用L16(45)正交实验，对苦瓜SSR-PCR 反应体系的Mg2+浓度、dNTPs浓度、模板DNA量、Taq聚合酶量、引物浓度等5个因素的4个水平进行优化试验，并通过正交设计直观分析法对Mg2+浓度、dNTPs浓度、Taq聚合酶量进行了单因素确定。结果表明：最佳反应体系为1 μL 10×PCR buffer，Mg2+浓度为1.75 mmol?L-1，dNTPs浓度为0.25 mmol?L-1，Taq DNA 聚合酶1 U，DNA 100 ng，引物0.20 mmol?L-1，ddH2O 补足至10 μL。
- Abstract:
- Taking Momordica charantia ‘MC9’ as test material,an orthogonal experiment design of L16(45) with four levels and five factors was carried out，the factors were the concentration of Mg2+，the concentration of dNTPs，Taq polymerase，DNA template and the concentration of primers.The concentration of Mg2+，the concentration of dNTPs and DNA polymerase using single factor experiment design were also determined.The results showed that the optimum system containing 1 μL 10×PCR buffer,Mg2+ 1.75 mmol?L-1，dNTPs 0.25 mmol?L-1，Taq DNA polymerase 1 U，DNA template 100 ng，primer 0.20 mmol?L-1.
参考文献/References:
[1]罗冉,吴委林,张旸,等.SSR分子标记在作物遗传育种中的应用[J].基因组学与应用生物学,2010(1):137-143. [2]徐彦军，刘洋.苦瓜外观性状AFLP反应体系的建立及优化[J].中国农学通报,2012,28(4):125-130. [3]郭大龙,吴正景,郑玉萍,等.苦瓜SRAP反应体系的建立与优化[J].安徽农业科学,2008,36(18):7583-7585. [4]刘萌芽,黄如葵,黄玉辉,等.正交直观分析法和新复极差法优化苦瓜SRAP反应体系研究[J].北方园艺,2014(5):85-88. [5]田丽波,谷幸幸,商桑,等.苦瓜基因组DNA的提取及ISSR扩增体系的优化[J].中国农学通报,2013，29(4):88-93. [6]张菊平,巩振辉,张长远,等.苦瓜RAPD分析体系的优化研究[J].河南农业大学学报,2003(1):49-53. [7]WANG S Z，PAN L，HU K，et al.Development and characterization of polymorphic microsatellite markers in Momordica charantia(Cucurbitaceous)[J].American Journal of Botany,2010，97(8)：75-78. [8]GUO D L，ZHANG J P，XUE Y M，et al.Isolation and characterization of 10 SSR markers of Momordica charantia（Cucurbitaceous）[J].American Journal of Botany，2012，99(5):182-183. [9]何正文,刘运生,陈立华,等.正交设计直观分析法优化PCR条件[J].湖南医科大学学报,1998(4):76-77. [10]孙佩光,奚如春,钮世辉,等.油茶SRAP-PCR反应体系的建立和优化[J].基因组学与应用生物学,2010(6):1192-1199. [11]周静,陈书霞,程智慧,等.大蒜SSR体系的建立与优化[J].西北农业学报,2011(11):117-122. [12]何仁锋,冯尚国,陈喆,等.药用菊花SSR-PCR反应体系优化及引物筛选[J].分子植物育种,2015(2):367-378. [13]徐阳,陈金慧,王颖,等.杉木SSR-PCR体系优化[J].林业科技开发,2014（1）:15-20.
相似文献/References:
[1]周金梅,赵文若,建德锋,等.不同品种和栽植密度及整枝方式对黄秋葵产量的影响[J].北方园艺,2013,37(04):34.
[2]刘晓莉,田海娟,钟容,等.盐沉淀法提取紫苏活性蛋白的工艺研究[J].北方园艺,2013,37(04):147.
[3]史艳财,邹 蓉,韦记青,等.黄花倒水莲总黄酮提取工艺研究[J].北方园艺,2014,38(12):134.
SHI Yan-cai,ZOU Rong,WEI Ji-qing,et al. Study on Extraction Condition of Flavonoid in Polygala fallax[J].Northern Horticulture,2014,38(10):134.
[4]叶 林,李春江,张光弟,等.不同育苗穴数、基质配比和施肥量对温室辣椒幼苗生长及其秧苗质量的影响[J].北方园艺,2014,38(13):50.
YE Lin,LI Chun-jiang,ZHANG Guang-di,et al.Effects of Different Seedling Number of Holes，Substrates and Fertilization on the Growth and Quality of Greenhouse Pepper Seedlings[J].Northern Horticulture,2014,38(10):50.
[5]张 颖.青蒿素的提取工艺优化研究[J].北方园艺,2014,38(11):122.
ZHANG Ying.Study on Optimization of the Extraction of Artemisinin[J].Northern Horticulture,2014,38(10):122.
[6]杨申明,王振吉,沈继森.彝药猪鬃草中总黄酮提取工艺研究[J].北方园艺,2014,38(11):125.
YANG Shen-ming,WANG Zhen-ji,SHEN Ji-sen.Study of the Extraction Technology of Total Flavonoids in Adiantum capillus-veneris[J].Northern Horticulture,2014,38(10):125.
[7]王 嵩,马 杰,赵 维,等.黔西北景天三七高产栽培优化研究[J].北方园艺,2014,38(11):147.
WANG Song,MA Jie,ZHAO Wei,et al.Research on Optimazation Growing Cutivation of Sedum aizoon L. in Northwest Guizhou[J].Northern Horticulture,2014,38(10):147.
[8]王亚红,王亚丽,祝 波.瓦松总黄酮的索氏提取工艺研究[J].北方园艺,2014,38(10):130.
WANG Ya-hong,WANG Ya-li,ZHU Bo.Study on Soxhlet Extraction Technology of Total Flavonoids From Orostachys fimbriatus[J].Northern Horticulture,2014,38(10):130.
[9]杨艳俊,王一洋.微波辅助法提取景天三七中齐墩果酸的工艺研究[J].北方园艺,2014,38(10):152.
YANG Yan-jun,WANG Yi-yang.Study on Extraction Technology of Oleanolic Acid From Sedum aizoon L.[J].Northern Horticulture,2014,38(10):152.
[10]姜 喜,陈加利,肖 巍,等.扁桃茎段增殖培养基的筛选[J].北方园艺,2013,37(22):111.
JIANG Xi,CHEN Jia-li,XIAO Wei,et al.Screening of the Multiplying Medium of Almond Stem[J].Northern Horticulture,2013,37(10):111.
备注/Memo
第一作者简介：王心迪（1991-），女，硕士研究生，研究方向为苦瓜核心种质筛选。E-mail：xindikaka@126.com. 责任作者：黄如葵（1969-），女，博士，研究员，现主要从事蔬菜育种及种子学等研究工作。E-mail：rkhuang@gxaas.net. 基金项目：现代农业产业技术体系建设专项资助项目(CARS-25);国家科技计划资助项目(2014BAD05B04);广西科学研究与技术开发计划资助项目(桂科合14125008-2-19)；广西农业科学院基本科研业务专项资助项目(2015YT66)。