[1]贾俊香,崔连伟,李娜,等.大葱ISSR反应体系的建立及优化[J].北方园艺,2017,41(12):92-96.[doi:10.11937/bfyy.201712023]
　JIA Junxiang,CUI Lianwei,LI Na,et al.Establishment and Optimalization of ISSR Reaction System on Welsh Onion[J].Northern Horticulture,2017,41(12):92-96.[doi:10.11937/bfyy.201712023]
点击复制

大葱ISSR反应体系的建立及优化

《北方园艺》[ISSN:1001-0009/CN:23-1247/S] 卷: 第41卷 期数: 2017年12 页码: 92-96 栏目: 出版日期: 2017-06-30
Title:
Establishment and Optimalization of ISSR Reaction System on Welsh Onion
作者:
贾俊香; 崔连伟; 李娜; 姜滨滨
（辽宁省农业科学院 蔬菜研究所，辽宁 沈阳 110161）
Author(s):
JIA Junxiang; CUI Lianwei; LI Na; JIANG Binbin
(Institute of Vegetables，Liaoning Academy of Agricultural Sciences，Shenyang，Liaoning 110161)
关键词:
大葱; ISSR; 反应体系; 单因素试验; 正交设计
Keywords:
Allium; ISSR; reaction system; single factor test; orthogonal design
DOI:
10.11937/bfyy.201712023
文献标志码:
A
摘要:
以大葱嫩叶为试材，采用单因素试验和正交实验探讨了Mg2+浓度、dNTPs浓度、引物浓度、Taq DNA聚合酶用量、模板DNA以及Buffer用量对大葱ISSR-PCR扩增的影响，建立了大葱ISSR反应体系。结果表明：优化后的最佳反应体系为10 μL反应体系中，10×Buffer缓冲液1.0 μL，Mg2+浓度为2.0 mmol?L-1，dNTPs浓度为0.3 mmol?L-1，引物浓度为0.65 μmol?L-1，Taq酶1.0 U，模板DNA为60 ng，用灭菌的超纯水补齐体积至10 μL。对大葱71份样品材料进行ISSR-PCR 扩增体系的检测结果显示，该优化体系扩增的产物条带清晰，稳定性高，为大葱种质ISSR分子标记遗传结构分析提供参考依据。
Abstract:
The young leaves of welsh onion was used as the test materials,and adopting single factor test and orthogonal test,the effect of Mg2+concentration,dNTPs dosage,primers amount,Taq DNA polymerase,template DNA and different buffer dosage on ISSR-PCR amplification of welsh onion were investigated，and the welsh onion ISSR-PCR reaction system was set up.The results showed that the optimized reaction system was established as follows,in the total reaction system with amount of 10 μL,10× buffer 1.0 μL,Mg2+ 2.0 mmol?L-1，dNTPs 0.3 mmol?L-1，primer 0.65 μmol?L-1，Taq DNA polymerase 1.0 U，template DNA 60 ng.The obtained ISSR-PCR amplification system in 71 materials was detected，and the results suggested that the bands were clear and with high stability，would provide the genetic basis for the further analysis of welsh onions.

参考文献/References:

 

[1]ZIETKIEWICZ E,RAFALSKI A,LABUDA D.Genomefingerprinting by simple sequence repeat(SSR)-anchored polymerase chain reaction amplification[J].Genonics,1994,20(2):176-183.

[2]奥斯伯Ｆ.分子生物学试验指南[M].北京:科学出版社,1998.

[3]张明,李廷龙.基于ISSR标记的韭菜种质资源遗传多样性初探[J].西北农业学报,2012,21(1):146-150.

[4]金雪,周新刚,刘守伟,等.48份分蘖洋葱种质资源遗传多样性的ISSR标记和农艺性状分析[J].中国蔬菜,2013(2):27-34.

[5]徐启江,崔成日,贾铁金.洋葱种质资源遗传多样性的ISSR分析[J].中国农学通报,2007,23(6):126-130.

[6]田艳伶,李志辉,杨模华,等.钩栗ISSR-PCR反应体系的建立与优化[J].中南林业科技大学学报,2015,35(2)32-36.

[7]廖安红,杨鑫,陈红.贵州桃种质资源ISSR标记体系建立及遗传多样性分析[J].分子植物育种,2015,13(12):2773-2781.

[8]郑姗,张立杰,谢丽雪,等.蓝莓品种ISSR指纹图谱构建的初步研究[J].福建农业学报,2014,29(12):1198-1201.

[9]马丽,侯乐峰,郝兆祥.82个石榴品种遗传多样性的ISSR分析[J].果树学报,2015,32(5):741-750.

[10]张冰雪,范付华,乔光,等.贵州地方火龙果芽变种质DNA指纹图谱及遗传多样性的ISSR分析[J].果树学报,2013(4):573-577.

[11]陈红,杨迤然.贵州李资源遗传多样性及亲缘关系的ISSR分析[J].果树学报,2014,31(2):175-180.

[12]张起,安华明.贵州砂梨种质资源遗传多样性的ISSR分析[J].果树学报,2015,32(1):6-12.

[13]史辉,董晓菲,阮少江,等.太子参ISSR反应体系的优化[J].安徽农业大学学报,2016,43(5):820-826.

[14]雒新艳,王晨,戴思兰,等.基于ISSR标记的大菊品种资源遗传多样性分析[J].中国农业科学,2013,46(11):2394-2402.

[15]张娜,李群,高兴旺,等.葡萄ISSR-PCR反应体系的建立与正交设计优化[J].江苏农业科学,2016,44(9):47-50.

[16]石琳,胡延萍,王建科,等.云生毛茛ISSR-PCR体系优化与引物筛选[J].生物技术通报,2016,32(9):65-71.

[17]JOSHI S P,GUPTA V S,AGGARWAL R K,et al.Genetic diversity and phylogenetic relationship as revealed by inter simple sequence repeat(ISSR)polymorphism in the genus Oryza[J].Theor Appl Genet,2000,100(8):1311-1320.

相似文献/References:

[1]肖志娟,翟梅枝,许静,等.不同核桃品种的ISSR标记分析[J].北方园艺,2013,37(04):103.
[2]刘志曦,李志强,魏海莲,等.平菇ISSR遗传多样性分析[J].北方园艺,2014,38(12):94.
　LIU Zhi-xi,LI Zhi-qiang,WEI Hai-lian,et al. Phylogenetic Analysis of Oyster Mushrooms by ISSR Marker[J].Northern Horticulture,2014,38(12):94.
[3]李 娜,张存旭,崔晓燕,等.栓皮栎ISSR反应体系的建立[J].北方园艺,2014,38(13):93.
　LI Na,ZHANG Cun-xu,CUI Xiao-yan,et al.Establishment of ISSR Reaction System in Quercus variabilis [J].Northern Horticulture,2014,38(12):93.
[4]王鸿磊,丁 强,吕 蔚,等.我国双孢菇菌株与国外引进菌株间亲缘关系的ISSR分析[J].北方园艺,2014,38(11):96.
　WANG Hong-lei,DING qiang,LV Wei,et al.Genetic Relationship Based on ISSR Markers Among Agaricus bisporus Strains From China and Overseas[J].Northern Horticulture,2014,38(12):96.
[5]李 娜,张存旭,刘 伟,等.毛梾ISSR-PCR反应体系的建立[J].北方园艺,2013,37(22):114.
　LI Na,ZHANG Cun-xu,LIU Wei,et al.Establishment of ISSRPCR Reaction System of Cornus walteri[J].Northern Horticulture,2013,37(12):114.
[6]黄伟,刘灵娣,王荣荣,等.冀西北坝上地区大葱引种试验[J].北方园艺,2013,37(10):42.
　HUANG Wei,LIU Ling-di,WANG Rong-rong,et al.Study on Comparative Experiment of Green Chinese Onion Varieties in the Northwest of Hebei Province[J].Northern Horticulture,2013,37(12):42.
[7]梁新安,邵秀丽,张慎璞,等.大葱标准化生产施肥配方技术研究[J].北方园艺,2013,37(12):172.
　LIANG Xin-an,SHAO Xiu-li,ZHANG Shen pu,et al.Study on the Fertilization Formula of Standardization Production Scallion[J].Northern Horticulture,2013,37(12):172.
[8]黄萌,陈尚钘,杨光耀,等.古樟ISSR扩增条件的优化[J].北方园艺,2013,37(07):116.
　HUANG Meng,CHEN Shang-xing,YANG Guang-yao,et al.Optimization of ISSR Amplification Conditions in Ancient Camphor-trees[J].Northern Horticulture,2013,37(12):116.
[9]吴春燕,高义,宋廷宇,等.正交设计优化大白菜ISSR-PCR 反应体系[J].北方园艺,2012,36(05):127.
　WU Chun-yan,GAO Yi,SONG Ting-yu,et al.Optimization of an ISSR-PCR Reaction System of Chinese Cabbage by Orthogonal Design[J].Northern Horticulture,2012,36(12):127.
[10]陈名红,李 玉,刘 多,等.适于ISSR-PCR扩增的百合基因组DNA的提取[J].北方园艺,2012,36(24):108.
　CHEN Ming-hong,LI Yu,LIU Duo,et al.Extraction of Genomic DNA for ISSR-PCR Reaction in Lilium[J].Northern Horticulture,2012,36(12):108.

备注/Memo

第一作者简介：贾俊香（1979-），女，硕士，助理研究员，现主要从事大葱新品种选育等研究工作。E-mail：xiangjj1129@163.com.基金项目：辽宁省科技厅自然科学基金资助项目（2014027019）。

更新日期/Last Update: 2017-06-27