| 利用Inverse PCR快速筛选单个T-DNA拷贝转基因水稻植株的方法 |
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| 引用本文： | 章冰,黄健秋,卫志明.利用Inverse PCR快速筛选单个T-DNA拷贝转基因水稻植株的方法[J].分子细胞生物学报,1999(2). |
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| 作者姓名： | 章冰 黄健秋 卫志明 |
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| 作者单位： | 中国科学院上海植物生理研究所，中国科学院上海植物生理研究所，中国科学院上海植物生理研究所 上海 200032，上海 200032，上海 200032 |
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| 摘 要： | 为了获得单个T-DNA插入拷贝的植株, 我们建立了一套利用Inverse PCR(IPCR)快速检测转基因水稻中T-DNA拷贝数的方法。用IPCR的方法可以扩增出与已知T-DNA序列相邻的水稻基因组DNA未知序列,由此推测转基因水稻植株中T-DNA的拷贝数。我们共对15个转化株系20棵不同植株的DNA进行了IPCR检测。其中12株表现为T-DNA单拷贝插入,3株为双拷贝插入,1株为三拷贝插入。另外4株未检测到T-DNA插入拷贝。IPCR分析结果经过Southern杂交和测序的验证。
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| 关 键 词： | Inverse PCR T-DNA拷贝数 水稻(Oryza sativa L.) 农杆菌介导转化 |
A QUICK METHOD TO ESTIMATE THE T-DNA COPY NUMBER IN TRANSGENIC RICE USING INVERSE PCR (IPCR) |
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| ZHANG Bing HUANG Jian Qiu WEI Zhi Ming.A QUICK METHOD TO ESTIMATE THE T-DNA COPY NUMBER IN TRANSGENIC RICE USING INVERSE PCR (IPCR)[J].Journal of Molecular Cell Biology,1999(2). |
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| Authors: | ZHANG Bing HUANG Jian Qiu WEI Zhi Ming |
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| Abstract: | In order to obtain single T-DNA copy trans-genie rice, we have established a quick method to estimate the T-DNA copy number in transgenic rice using inverse PCR (IPCR). IPCR was used to amplify junction fragments, i. e. plant genom-ic DNA sequences flanking the known T-DNA sequences, which will help to estimate the T-DNA copy number in transgenic rice. We have
analyzed 20 transgenic plants of 15 transgenic lines. Most plants (12) contain one integrated T-DNA copy per genome, 3 plants contain two and 1 plant contains 3 copies. In 4 transgenic plants no T-DNA copies could be detected using this method. The IPCR results were further tested by Southern analysis and sequence analysis. |
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| Keywords: | Inverse PCR T-DNA copy number Rice (Oryza sativa) Agrobacterium-medinted transformation |
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