ACC合酶cDNA克隆及其对美洲黑杨体内乙烯产生的反义抑制

林业科学 ›› 1999, Vol. 35 ›› Issue (3): 10-15.

ACC合酶cDNA克隆及其对美洲黑杨体内乙烯产生的反义抑制

李明亮¹,韩一凡¹,邱德有¹,李凝²,田颖川³

1. 中国林业科学研究院林业研究所北京 100091
2. 香港科技大学生物学系香港
3. 中国科学院微生物研究所北京 100080

收稿日期:1998-08-12 出版日期:1999-05-25 发布日期:1999-05-25
基金资助:
国家自然科学基金项目“反义ACC合酶和ACC氧化酶基因对杨树生长发育的影响”(39500119)

CLONING OF ACC SYNTHASE cDNA AND ITS INHIBITION OF ETHYLENE SYNTHESIS BY ITS ANTISENSE RNA IN TRANSGENIC POPULUS DELTOIDES

Mingliang Li¹,Yifan Han¹,Deyou Qiu¹,Ning Li²,Yingchuan Tian³

1. The Institute of Forestry, Chinese Academy of Forestry Beijing 100091
2. Department of Biology, the Hong Kong University of Science and Technology Hong Kong
3. The Institute of Microbiology, Chinese Academy of Sciences Beijing 100080

Received:1998-08-12 Online:1999-05-25 Published:1999-05-25

摘要/Abstract

摘要：

利用RT-PCR技术克隆了大豆ACC合酶基因GMCACCSI编码区1.5kb的cDNA片段,经酶切图谱分析和序列分析鉴定后,反向插入到2元载体pBin438中,构建了表达ACC合酶反义RNA的植物表达载体,转化农杆菌后用该农杆菌侵染美洲黑杨叶片,在含卡那霉素的MS培养基上选择转化子和植株再生,通过PCR检测从抗卡那植株中选到15株转基因植株, Southern杂交分析初步确证了外源基因是以单拷贝插入到杨树基因组DNA中;对杨树幼苗乙烯释放的测定结果表明转基因杨树幼苗的乙烯释放量为对照的22%。

关键词: ACC(l-aminocyclopropane-l-carboxylate)合酶, 基因, 反义RNA, 美洲黑杨

Abstract:

A 1.5 kb fragment of ACC synthase cDNA fragment prepared from total RNA of soybean was amplified by polymerase chain reaction (PCR) and cloned into pGEM^®-T vector. The cloned ACC synthase gene was further inserted into a binary vector, pBin438, in an inverted orientation between the CaMV 35S promoter and Nos 3'termination sequence (pBACS). Transgenic poplar plants were obtained by regeneration of Agrobacterium-mediated transformation of leaves of Populus deltoides. PCR and Southern blotting analyses confirmed the integration of a single antisense ACC synthase gene in transformed poplar genome. The results form reverse transcription PCR (RT-PCR) of RNAs isolated from transgenic poplar leaves confimed that the antisense RNA of ACC synthase presented in these transgenic plants. The amount of ethylene released from transgenic poplar was reduced significantly to about 22% of that released from non-transformed control poplars.

Key words: ACC(l-aminocyclopropane-l-carboxylate)synthase, Gene, Antisense RNA, Populus deltoides

李明亮,韩一凡,邱德有,李凝,田颖川. ACC合酶cDNA克隆及其对美洲黑杨体内乙烯产生的反义抑制[J]. 林业科学, 1999, 35(3): 10-15.

Mingliang Li,Yifan Han,Deyou Qiu,Ning Li,Yingchuan Tian. CLONING OF ACC SYNTHASE cDNA AND ITS INHIBITION OF ETHYLENE SYNTHESIS BY ITS ANTISENSE RNA IN TRANSGENIC POPULUS DELTOIDES[J]. Scientia Silvae Sinicae, 1999, 35(3): 10-15.

图/表 5

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