Scientia Agricultura Sinica ›› 2013, Vol. 46 ›› Issue (17): 3635-3642.doi: 10.3864/j.issn.0578-1752.2013.17.012

• HORTICULTURE • Previous Articles     Next Articles

Sequence Analysis and Prokaryotic Expression of Two Ethylene Responsive Factors MdAP2D4 and MdAP2D19

 TIAN  Yi-12, CHEN  Ke-Qin-1, AN  Xiu-Hong-1, HAO  Hong-Mei-2, LIU  Zhi-3, CONG  Pei-Hua-2, HAO  Yu-Jin-1   

  1. 1.College of Horticulture Science and Engineering/State Key Laboratory of Crop Biology, Taian 271018, Shandong
    2.Institute of Pomology, Chinese Academy of Agricultural Sciences/Key Laboratory of Fruit Germplasm Resources Utilization, Ministry of Agriculture, Xingcheng 125100, Liaoning
    3.Liaoning Institute of Pomology, Xiongyue 115009, Liaoning
  • Received:2013-03-11 Online:2013-09-01 Published:2013-07-12

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Abstract: 【Objective】 Cloning, sequence and expression analysis of two ERF (ethylene responsive factor) transcription factors from Malus domestica Borkh and expression of their fusion proteins in E. coli were determined to identify their disease resistant function and molecular mechanism.【Method】ESTs derived from suppression subtractive hybridization were blasted, and two EFR genes, MdAP2D4 and MdAP2D19, were cloned which contain these ESTs; Subsequently, phylogenetic relationship including the two apple proteins and Arabidopsis AP2 domain-containing proteins was analyzed using MEGA4.1 and the multiple alignment of AP2/ERF domain was constructed; Additionaly, the in vitro shoot cultures of ‘Gala’ apple were used to identify the two genes expression in response to exogenous MeJA; Finally, the two genes were inserted into vector pGEX-4T-1, and IPTG was used to induce the fusion proteins expression in E. coli BL21.【Result】The phylogenetic relationship of MdAP2D4 and MdAP2D19 was closer with Arabidopsis ERF proteins of group B3 than others. The transcript levels of MdAP2D4 and MdAP2D19 were higher in leaves than other organs and could be induced by exogenous MeJA. The results of SDS-PAGE demonstrated that the fusion proteins of the two genes could express in E.coli BL21 induced by IPTG.【Conclusion】 MdAP2D4 and MdAP2D19 are ERF transcription factors belonging to group B3, and they can be induced by exogenous MeJA.

Key words: suppression subtractive hybridization , Methyl jasmonate , ethylene responsive factor , prokaryotic expression , disease resistance

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