Characterization of an ethylene-induced esterase gene isolated from Citrus sinensis by competitive hybridization.
Characterization of an ethylene-induced esterase gene isolated from Citrus sinensis by competitive hybridization.
复制标题
通过竞争杂交从柑橘中分离乙烯诱导的酯酶基因的表征。
DOI：
10.1034/j.1399-3054.2001.1130215.x
复制
发表时间：
2001
期刊：
影响因子：
--
通讯作者：
D. Holland
中科院分区：
文献类型：
--
作者：
G. Zhong;R. Goren;J. Riov;E. Sisler;D. Holland
利用一种简单的新方法--竞争杂交法，分离了乙烯诱导的柑橘（Citrus sinensis [L.]奥斯贝克湾Shamarin）leaves.其中一个分离的基因，乙烯诱导的酯酶基因（EIE），进一步的特点。该基因的推导蛋白质序列与已知参与次级代谢的几种植物α/β水解酶基因家族成员的蛋白质序列相似。北方印迹分析表明，EIE mRNA在乙烯处理后4 h内即被诱导表达，24 h后EIE mRNA积累到很高水平。乙烯对EIE的诱导作用可被1-甲基环丙烯（一种有效的乙烯感知抑制剂）所抵消，表明EIE的表达是乙烯依赖的。EIE的细菌表达蛋白被针对Pir 7 b的抗血清所识别，Pir 7 b是一种由非宿主病原体假单胞菌pv.你好使用抗Pir 7 b抗体在乙烯处理的叶中鉴定EIE蛋白。α-醋酸萘酯酶的积累伴随着乙烯处理的柑橘叶片中EIE蛋白的增加。酶活性测定和Western分析证实该酯酶为EIE。
A simple new method, competitive hybridization, for identification of differentially regulated genes was used to isolate novel genes induced by ethylene in citrus (Citrus sinensis [L.] Osbeck cv. Shamouti) leaves. One of the isolated genes, an ethylene-induced esterase gene (EIE), was further characterized. The deduced protein sequence of this gene shows a similarity to those of several plant alpha/beta hydrolase gene family members, which are known to be involved in secondary metabolism. Northern blot analysis demonstrated that EIE mRNA was induced by ethylene within 4 h and accumulated to a very high level 24 h after the initiation of ethylene treatment. Induction of EIE by ethylene could be counteracted by 1-methylcyclopropene, a potent ethylene perception inhibitor, indicating that the expression of EIE is ethylene-dependent. The bacterially expressed protein of EIE was recognized by antiserum against Pir7b, a naphthol AS esterase induced in rice by the non-host pathogen, Pseudomonas syringae pv. syringae. The EIE protein was identified in ethylene-treated leaves using anti-Pir7b antibodies. An alpha-naphthyl acetate esterase accumulated concomitantly with the increase in EIE protein in ethylene-treated citrus leaves. An enzyme activity assay followed by western analysis confirmed that the esterase was EIE.