[1]宁爱玲,杜海平,喻德跃,等.GmAOC3-基因转化载体构建及转化大豆的初步研究[J].大豆科学,2015,34(04):588-596.[doi:10.11861/j.issn.1000-9841.2015.04.0588]
　NING Ai-ling,DU Hai-ping,YU De-yue,et al.Construction of Transformation Vector of GmAOC3 Gene and Preliminary Study on the Transformation of Soybean[J].Soybean Science,2015,34(04):588-596.[doi:10.11861/j.issn.1000-9841.2015.04.0588]
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GmAOC3-基因转化载体构建及转化大豆的初步研究

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第34卷 期数: 2015年04期 页码: 588-596 栏目: 出版日期: 2015-08-25
Title:
Construction of Transformation Vector of GmAOC3 Gene and Preliminary Study on the Transformation of Soybean
作者:
宁爱玲; 杜海平; 喻德跃; 王慧
南京农业大学 大豆研究所/国家大豆改良中心/农业部大豆生物学与遗传育种重点实验室/作物遗传与种质创新国家重点实验室，江苏 南京 210095
Author(s):
NING Ai-ling;  DU Hai-ping;  YU De-yue;  WANG Hui
Soybean Research Institute, Nanjing Agricultural University/National Center for Soybean Improvement, Ministry of Agriculture/Key Laboratory for Biology and Genetic Improvement of Soybean (General), Ministry of Agriculture / National Key Laboratory of Crop Genetics and Germplasm Enhancement; Nanjing 210095,China.
关键词:
大豆; 丙二烯氧化物环化酶; 载体构建; 遗传转化; 分子检测
Keywords:
Soybean;  Allene oxide cyclase;  Vector construction;  Genetic transformation;  Molecular detection
DOI:
10.11861/j.issn.1000-9841.2015.04.0588
文献标志码:
A
摘要:
选用内切酶SacⅠ和MluⅠ切割目标基因和载体，构建了携带-bar-基因的重组载体pBA002-GmAOC3，选用2个大豆品种（Jack和南农88-1）和2种外植体（子叶节和整个子叶节），研究大豆品种和外植体对根癌农杆菌介导的子叶节转化-GmAOC3-基因的影响。结果表明：外植体为整个子叶节的大豆品种Jack的出芽率和转化率最高，分别为79.5%和2.27%。经QuickStix PAT/bar-基因试纸条检测、目标基因的分子检测和草丁膦抗性鉴定，共得到转-GmAOC3-基因的T0代阳性株系3株，T1代阳性转基因大豆6株，其中5株以Jack品种为受体的转基因大豆-GmAOC3-基因的表达量均显著高于非转基因植株，荧光定量拷贝数检测结果显示，4株转基因单株为单拷贝，2株为双拷贝。
Abstract:
Allene oxide cyclase (AOC) plays a key role in the synthesis of jasmonic acid, which is an important defense signaling molecular in plant. In this study, we constructed a recombinant vector pBA002-GmAOC3 with bar gene using double enzymes cleavage method (SacⅠand MluⅠ) and the recombinant vector was successfully transformed into soybean via Agrobacterium transformation system. Two soybean varieties (Jack and Nannong 88.1) and explants (cotyledonary node and whole cotyledonary node) were used to study their effects on the Agrobacterium-mediated transformation of GmAOC3. The results showed that the whole cotyledonary node of Jack had the highest budding rate (79.5%) and conversion rate (2.27%) Based on the results of QuickStix PAT/bar- test paper detection, molecular detection of target DNA and Basta resistance test of leaves, we obtained three T0generation GmAOC3 positive transgenic soybean plants and six T1generation GmAOC3 positive transgenic soybean plants, of which five T1generation transgenic plants from Jack showed higher expression level of GmAOC3 than Jack. We also used the fluorescence quantitative PCR to detect the copy number of target gene Besides two T1generation transgenic plants with two copies of bar genes, the others contained one copy of bar gene. The data in this study would give some references to soybean genetic transformation and the GmAOC3 transgenic plant obtained in this study could be used in soybean insect resistance breeding in future.

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备注/Memo

基金项目：国家自然科学基金（31201230）；江苏省自然科学基金（BK2012768）。

第一作者简介：宁爱玲（1987-），女，硕士，主要从事大豆基因工程研究。E-mail:futurena1@163.com。
通讯作者：王慧（1977-），女，博士，副教授，主要从事大豆抗虫研究。E-mail:wanghuio@njau.edu.cn。

更新日期/Last Update: 2015-08-31