| 以小麦根尖分生组织细胞核及染色体制备高分子量DNA |
| |
| 引用本文： | 郭东伟,佘茂云,李连城,陈耀峰,陈明,徐兆师,马有志. 以小麦根尖分生组织细胞核及染色体制备高分子量DNA[J]. 作物学报, 2006, 32(12): 1920-1923 |
| |
| 作者姓名： | 郭东伟 佘茂云 李连城 陈耀峰 陈明 徐兆师 马有志 |
| |
| 作者单位： | 1.西北农林科技大学农学院，陕西杨凌712100；2中国农业科学院作物科学研究所/农业部遗传育种重点开放实验室，北京100081；3新疆农业大学农学院，新疆乌鲁木齐830052 |
| |
| 基金项目： | 西北农林科技大学校科研和教改项目 |
| |
| 摘 要： | 为了克服以叶片为材提取HMW DNA的局限性，优化了一套简便、实用的提取方法。该法以细胞周期同步化处理的根尖分生组织为材，利用机械匀浆释放细胞核或中期染色体制备悬浮液，再以蔗糖密度梯度离心和流式分选技术分别分离细胞核和染色体，经去蛋白、酶切，透析获得HMW DNA。经检测，来自200条根尖（10个胶块）的HMW DNA的浓度约为4~20 ng/µL，而连接、转化后的BAC克隆平均插入片段超过100 kb。证明该法适用于提取HMW DNA以构建BAC文库。
|
| 关 键 词： | 高分子量DNA 蔗糖密度梯度离心 细胞周期同步化 染色体流式分选 BAC |
| 收稿时间： | 2006-03-10 |
| 修稿时间： | 2006-03-10 |
High Molecular Weight DNA Preparation from Nuclei or Chromosomes of Root Meristem of Wheat |
| |
| GUO Dong-Wei,SHE Mao-Yun,LI Lian-Cheng,CHEN Yao-Feng,CHEN Ming,XU Zhao-Shi,MA You-Zhi. High Molecular Weight DNA Preparation from Nuclei or Chromosomes of Root Meristem of Wheat[J]. Acta Agronomica Sinica, 2006, 32(12): 1920-1923 |
| |
| Authors: | GUO Dong-Wei SHE Mao-Yun LI Lian-Cheng CHEN Yao-Feng CHEN Ming XU Zhao-Shi MA You-Zhi |
| |
| Affiliation: | 1.Agronomy College, Northwest Sci-Tech University of Agriculture and Forestry, Yangling 712100, Shaanxi;2.Key Laboratory of Crop Genetics and Breeding, Ministry of Agriculture/ Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081;3.Agronomy College, Xinjiang Agricultural University, Urumqi 810012, Xinjiang, China |
| |
| Abstract: | A simple and practical method was optimized to overcome the limitations of current method by which HMW DNA was prepared from leaf powder. Root tips of synchronized meristem were homogenized to make preparing suspension containing intact metaphase chromosomes and nuclei, which were further isolated by sucrose gradient centrifugation and flow sorting, respectively. Finally, HMW DNA was prepared with deproteinization, digestion and dialysis of nuclei and chromosomes. The results showed that concentration of the HMW DNA from 200 root tips (10 plugs) was about 4–20 ng/µL (4 ng/µL in chromosomes DNA and 20 ng/µL in nuclei DNA). The average insert fragment size was more than 100 kb after linage, transformation and insert fragment examination by pulse field gel electrophoresis for HMW DNA from nuclei. The similar results were obtained by HMW DNA prepared from chromosomes. It is confirmed that the method is applicable to prepare HMW DNA for construction of BAC library. |
| |
| Keywords: | BAC |
| 本文献已被 CNKI 维普 万方数据 等数据库收录！ |
| 点击此处可从《作物学报》浏览原始摘要信息 |
|
点击此处可从《作物学报》下载全文 |
|
