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园艺学报 ›› 2010, Vol. 37 ›› Issue (7): 1132-1138.

• 观赏植物 • 上一篇    下一篇

切花月季ACC合成酶基因的克隆和原核表达

谭远军1,薛璟祺2,仝 征1,马 男1,高俊平1,*   

  1. (1中国农业大学观赏园艺与园林系,北京100193;2中国农业科学院蔬菜花卉研究所,北京100081)
  • 收稿日期:2010-06-15 修回日期:2010-06-22 出版日期:2010-07-25 发布日期:2010-07-25
  • 通讯作者: 高俊平

Cloning and Recombinant Protein Expression of ACC Synthase Genes in Cut Roses(Rosa hybrida)

TAN Yuan-jun1,XUE Jing-qi2,TONG Zheng1,MA Nan1,and GAO Jun-ping1,*

  

  1. (1Department of Ornamental Horticulture and Landscape Architecture,China Agricultural University,Beijing 100193,China;2Institute of Vegetables and Flowers,Chinese Academy of Agricultural Sciences,Beijing 100081,China)
  • Received:2010-06-15 Revised:2010-06-22 Online:2010-07-25 Published:2010-07-25
  • Contact: GAO Jun-ping

摘要: 克隆了与伤害诱导相关的Rh-ACS1和与花朵开放进程及乙烯诱导相关的Rh-ACS3基因全长。结果表明,Rh-ACS1基因全长为2 132 bp,开放阅读框(ORF)长度为1 476 bp,推定编码491个氨基酸。Rh-ACS3基因的全长1 734 bp,ORF长度为1 545 bp,推定编码514个氨基酸。对这两个基因编码的蛋白进行原核表达分析,结果显示,在37 ℃条件下,0.6 mmol · L-1 IPTG诱导3 h后,携带Rh-ACS1 ORF和Rh-ACS3 ORF的原核表达载体分别在大肠杆菌中诱导生成了大量分子量约为70和60 kD的蛋白质,其大小与根据基因序列预测的理论值相一致。

关键词: 月季切花, ACS, 基因克隆, 原核表达

Abstract: Our previous work showed that the expression of ACC synthase genes was contributed to the effects of ethylene on hastening flower opening. In the present work,we isolated the wound-inducible and ethylene up-regulated ACS genes,namely Rh-ACS1 and Rh-ACS3,from cut rose petals. The full length of Rh-ACS1 is 2 132 bp,contains a 1 476 bp ORF which encodes a putative ACS protein with 491 aa,while the entire Rh-ACS3 is 1 734 bp in length,and contains a 1 545 bp ORF which encodes a putative ACS protein with 514 aa. The ORFs of Rh-ACS1 and Rh-ACS3 were inserted into pGEX-4T-1 and pET-30a to express the recombinant proteins in E.coli,respectively. The expression of predicted 70 kD and 60 kD recombinant proteins were induced by isopropyl β-D-thiogalacto-pyranoside(IPTG)at a final concentration of 0.6 mmol · L-1 for 3 h at 37 ℃.

Key words: cut rose, ACS, gene cloning, procaryotic expression

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