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中国农学通报

中国农学通报 ›› 2011, Vol. 27 ›› Issue (18): 218-222.

所属专题： 生物技术

• 生物技术科学 • 上一篇    下一篇

含BADH和Bar基因的植物表达载体的构建

李虹章 朱丹华 黄英运 郁晓敏 董德坤   

  • 收稿日期:2011-04-06 修回日期:2011-04-26 出版日期:2011-07-25 发布日期:2011-07-25
  • 基金资助:

    转基因生物新品种培育重大专项

Construction of a Plant Expression Vector Containing the BADH and Bar Genes

  • Received:2011-04-06 Revised:2011-04-26 Online:2011-07-25 Published:2011-07-25

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摘要：

采用PCR技术扩增出甜菜碱醛脱氢酶基因BADH，并于上下游引物5’端分别添加Xhol和SacI酶切位点和保护性碱基序列；TA克隆PCR产物至pTA2并进行测序验证；双酶切经过测序验证的重组质粒pTA2-BADH和植物表达载体pBA002，分别回收目的基因和pBA002载体片段，并进行连接转化，提取阳性质粒，然后进行双酶切、PCR扩增和进一步的测序验证。结果表明BADH基因已被完整、正确的插入到pBA002载体中，成功构建了含BADH和Bar基因的植物表达载体，为进一步的植物遗传转化奠定了基础。

关键词: 白点鲑, 白点鲑, 耗氧率, 窒息点

Abstract:

BADH (betaine-aldyhyde dehydrogenase, EC 1.2.1.8) cDNA was amplified by PCR, including XhoI (upstream) and SacI (downstream) restriction sites respectively. This BADH fragment was cloned into the vector pTA2 (obtaining pTA2-BADH) and verified by sequencing. Then the fragment was cloned from recombinant plasmid pTA2-BADH into the vector pBA002 by digestion and ligation. The obtained pBA002-BADH vector was further verified by enzyme digestion, PCR amplification, and sequencing. Using the plant expression vector pBA002-BADH, we are able to conduct further plant genetic transformation.