毛竹大片段双元细菌人工染色体基因组文库的构建

管雨; 杨洋; 张智俊; 罗淑萍; 汤定钦

doi:10.11833/j.issn.2095-0756.2011.04.001

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毛竹大片段双元细菌人工染色体基因组文库的构建

管雨 , 杨洋 , 张智俊 , 罗淑萍 , 汤定钦

文章导航 > 浙江农林大学学报 > 2011 > 28(4): 527-532

管雨, 杨洋, 张智俊, 罗淑萍, 汤定钦. 毛竹大片段双元细菌人工染色体基因组文库的构建[J]. 浙江农林大学学报, 2011, 28(4): 527-532. doi: 10.11833/j.issn.2095-0756.2011.04.001

引用本文:

GUAN Yu, YANG Yang, ZHANG Zhi-jun, LUO Shu-ping, TANG Ding-qin. Construction of a large genomic DNA fragments， BIBAC library for Phyllostachys pubescens[J]. Journal of Zhejiang A&F University, 2011, 28(4): 527-532. doi: 10.11833/j.issn.2095-0756.2011.04.001

Citation:

GUAN Yu, YANG Yang, ZHANG Zhi-jun, LUO Shu-ping, TANG Ding-qin. Construction of a large genomic DNA fragments， BIBAC library for Phyllostachys pubescens[J]. Journal of Zhejiang A&F University, 2011, 28(4): 527-532. doi: 10.11833/j.issn.2095-0756.2011.04.001

毛竹大片段双元细菌人工染色体基因组文库的构建

doi: 10.11833/j.issn.2095-0756.2011.04.001

1.
浙江农林大学亚热带森林培育国家重点实验室培育基地，浙江临安 311300;
2.
新疆农业大学农学院，新疆乌鲁木齐 830052

详细信息

通信作者: 张智俊

Construction of a large genomic DNA fragments， BIBAC library for Phyllostachys pubescens

1.
The Nurturing Station for the State Key Laboratory of Subtropical Silviculture，Zhejiang A & F University，Lin’an 311300，Zhejiang，China;
2.
College of Agronomy，Xinjiang Agricultural University，Urumqi 830052，Xinjiang，China

More Information

Corresponding author: ZHANG Zhi-jun

摘要: 从毛竹Phyllostachys pubescens幼嫩叶片中提取纯化得到高质量的基因组DNA，经限制性内切酶BamH I对它们进行梯度酶切，脉冲场电泳选择合适酶切DNA片段，与脱磷酸化处理过的质粒载体pCLD04541按质量比3∶1相互连接，转化大肠杆菌Escherichia coli DH10B感受态细胞进行蓝白斑筛选，挑取白色克隆，获得重组率较高的阳性克隆，构建了含有104个克隆的双元细菌人工染色体（BIBAC）基因组文库，并通过脉冲场电泳检测分析后确定，所构建的毛竹BIBAC文库平均插入片段为105 kb，约覆盖5倍毛竹基因组。该文库的构建为毛竹基因组研究做好了前期的基础工作。图8参18
- 分子生物学 /
- 毛竹 /
- 双元细菌人工染色体 /
- 基因组文库
Abstract: One plant-transformation-competent binary bacterial artificial chromosome（BIBAC） library was constructed which represents the first large genomic DNA fragment library generated for Phyllostachys pubescens. High-quality，genomic DNA extracted from young leaves of Phyllostachys pubescens was gradiently enzyme-digested with a gradient using BamH I. Desirable DNA fragments were isolated by pulsed field gel electrophoresis，ligated to the dephosphorylatedion carrier pCLD04541 with a mass ratio of 3∶1，and then transformed to Escherichia coli DH10B competent cells. This was followed by blue-white screening with establishment of a binary bacterial artificial chromosome（BIBAC） genome library. Results showed a high recombination positive colony from which a BIBAC genome library，consisting of 104 clones with an average insert fragment size of about 105 kb after detection with a pulsed field gel electrophoresis, was constructed. The BIBAC library was 5 times larger than the Phyllostachys pubescens genome. The construction of this BIBAC genome library laid a good foundation for related genome research. ［Ch，8 fig. 18 ref.］
- molecular biology /
- Phyllostachys pubescens /
- binary bacterial artificial chromosome （BIBAC） /
- genome library

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毛竹大片段双元细菌人工染色体基因组文库的构建

doi: 10.11833/j.issn.2095-0756.2011.04.001

1. 浙江农林大学亚热带森林培育国家重点实验室培育基地，浙江临安 311300;

2. 新疆农业大学农学院，新疆乌鲁木齐 830052

通信作者: 张智俊

刊出日期: 2011-08-20

关键词:

摘要: 从毛竹Phyllostachys pubescens幼嫩叶片中提取纯化得到高质量的基因组DNA，经限制性内切酶BamH I对它们进行梯度酶切，脉冲场电泳选择合适酶切DNA片段，与脱磷酸化处理过的质粒载体pCLD04541按质量比3∶1相互连接，转化大肠杆菌Escherichia coli DH10B感受态细胞进行蓝白斑筛选，挑取白色克隆，获得重组率较高的阳性克隆，构建了含有104个克隆的双元细菌人工染色体（BIBAC）基因组文库，并通过脉冲场电泳检测分析后确定，所构建的毛竹BIBAC文库平均插入片段为105 kb，约覆盖5倍毛竹基因组。该文库的构建为毛竹基因组研究做好了前期的基础工作。图8参18

English Abstract

Construction of a large genomic DNA fragments， BIBAC library for Phyllostachys pubescens

1. The Nurturing Station for the State Key Laboratory of Subtropical Silviculture，Zhejiang A & F University，Lin’an 311300，Zhejiang，China;

2. College of Agronomy，Xinjiang Agricultural University，Urumqi 830052，Xinjiang，China

Corresponding author: ZHANG Zhi-jun

Publish Date: 2011-08-20

Keywords:

molecular biology /
Phyllostachys pubescens /
binary bacterial artificial chromosome （BIBAC） /
genome library

Abstract: One plant-transformation-competent binary bacterial artificial chromosome（BIBAC） library was constructed which represents the first large genomic DNA fragment library generated for Phyllostachys pubescens. High-quality，genomic DNA extracted from young leaves of Phyllostachys pubescens was gradiently enzyme-digested with a gradient using BamH I. Desirable DNA fragments were isolated by pulsed field gel electrophoresis，ligated to the dephosphorylatedion carrier pCLD04541 with a mass ratio of 3∶1，and then transformed to Escherichia coli DH10B competent cells. This was followed by blue-white screening with establishment of a binary bacterial artificial chromosome（BIBAC） genome library. Results showed a high recombination positive colony from which a BIBAC genome library，consisting of 104 clones with an average insert fragment size of about 105 kb after detection with a pulsed field gel electrophoresis, was constructed. The BIBAC library was 5 times larger than the Phyllostachys pubescens genome. The construction of this BIBAC genome library laid a good foundation for related genome research. ［Ch，8 fig. 18 ref.］

引用本文: