| 深海细菌Pseudomonas sp.bIp-2黑色素生成条件研究及其hppD基因的克隆 |
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| 引用本文： | 韦海镧,于丽波,易志伟,汤熙翔.深海细菌Pseudomonas sp.bIp-2黑色素生成条件研究及其hppD基因的克隆[J].台湾海峡,2014(4):499-507. |
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| 作者姓名： | 韦海镧 于丽波 易志伟 汤熙翔 |
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| 作者单位： | 国家海洋局第三海洋研究所 国家海洋局海洋生物遗传资源重点实验室,福建 厦门,361005 |
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| 基金项目： | 国家自然科学基金资助项目 |
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| 摘 要： | 从大西洋深海沉积物中分离得到1株产黑色素细菌b Ip-2,16S rRNA基因序列分析表明该菌隶属于假单胞菌属（Pseudomonas）,与施氏假单胞菌（P.stutzeri）有最高的16S rRNA基因序列相似性（99%）.进一步对该菌的生长及产黑色素条件进行了初步研究,结果表明b Ip-2在稳定生长期后期才开始大量积累黑色素;b Ip-2可在4～37℃的温度范围下生长,最适生长温度为37℃,在28℃具有最高的黑色素产量;b Ip-2生长的pH范围偏碱性,在p H=7的最适生长条件下黑色素产量最高,同时碱性环境有利于黑色素的快速形成;b Ip-2可以在1%～9%的NaCl浓度范围中生长,在5%的NaCl浓度下具有最高的黑色素产量;低浓度的Fe^2＋（0.05 mmol/dm^3）可以提高黑色素的产量,Fe^2＋浓度≥0.20 mmol/dm^3时明显抑制黑色素的生成,而Fe^2＋浓度为1.00 mmol/dm^3时可完全抑制菌株的生长;低浓度的Cu^2＋（0.1 mmol/dm^3）对黑色素的产量无明显影响,Cu^2＋浓度≥0.5mmol/dm^3时可抑制黑色素的生成,而Cu^2＋浓度超过1.0 mmol/dm^3时可完全抑制菌株的生长.通过PCR扩增获得菌株b Ip-2的对羟苯基丙酮酸双加氧酶（hydroxyphenylpyruvic acid dioxygenase,HPPD）全长基因（1087 bp）,分析发现其编码的HPPD蛋白序列（含361氨基酸）与来自菌株Pseudomonas stutzeri RCH2的HPPD蛋白序列具有最高的同源性（序列相似性为97%）.这些结果为深入研究该细菌所产黑色素的合成,应用及生态作用奠定了基础.
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| 关 键 词： | 海洋生物学 黑色素 类黑素 对羟苯基丙酮酸双加氧酶 假单胞菌 |
Studies on the melanin production and gene cloning of hppD from the deep-sea bacterium Pseudomonas sp.bIp-2 |
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| WEI Hai-lan,YU Li-bo,YI Zhi-wei,TANG Xi-xiang.Studies on the melanin production and gene cloning of hppD from the deep-sea bacterium Pseudomonas sp.bIp-2[J].Journal of Oceanography In Taiwan Strait,2014(4):499-507. |
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| Authors: | WEI Hai-lan YU Li-bo YI Zhi-wei TANG Xi-xiang |
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| Institution: | (Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, SOA, Xiamen 361005, China) |
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| Abstract: | A melanin-producing strain was isolated from deep-sea sediment of the Atlantic Ocean and identified as Pseudomonas sp.b Ip-2 by 16 S rRNA gene sequencing.The study of growth and melanin production with varied culture conditions show that the melanin production in strain b Ip-2 is growth-associated.The maximum yield of pigment was obtained after the culture attained the stationary growth phase.The temperature range for growth is 4 ～37℃ with optimal growth at 37℃,and the maximum melanin production was observed at 28℃.The pH range for growth is 6.0 ～ 9.0,with both optimal growth and melanin formation both being at pH 7.0.Strain b Ip-2 grew in1% ～ 9% NaCl with a maximum melanin production at 5% NaCl.Low concentration of Fe（II）（0.05 mmol/dm^3）can improve the production of melanin.Fe^2＋of more than 0.20 mmol/dm^3 suppressed the melanin production and Fe^2＋of more than 1.00 mmol/dm^3 completely inhibited the growth of the strain.Cu^2＋of 0.1 mmol/dm^3 had no effect to the melanin production and Cu^2＋ of more than 0.5 mmol/dm^3 inhibited the formation of melanin.Cu^2＋over 1.0 mmol/dm^3 completely inhibited the growth of the strain.A 1 087 bp fragment of the full-length hpp D gene was amplified from the strain b Ip-2.The hpp D gene encoded an amino acid sequence sharing a 97% similarity with the HPPD from Pseudomonas stutzeri RCH2.These results laid a foundation to the study of the synthesis,application and ecological effect of the melanin from Pseudomonas sp.bIp-2. |
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| Keywords: | marine biology melanin pyomelanin hydroxyphenylpyruvic acid dioxygenase (HPPD) Pseudo-monas |
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