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CDS cloning and sequence analysis of neuronal nitric oxide synthase (nNOS)from plateau zokor (Myospalax baileyi)

ZHANG Xuze,XIE Ling,GUO Xinyi,CHEN Guihua,LIN Gonghua,DU Yurong,PANG Bo,GUO Songchang   

  • Online:2014-02-21 Published:2014-02-21


张湑泽 谢玲 郭新异 陈桂华 林恭华 都玉蓉 庞礴 郭松长   

  1. 中国科学院西北高原生物研究所

Abstract: Enduring hypoxia stress related to high altitude and subterranean burrows,the plateau zokor (Myospalax baileyi) has high efficiency in oxygen utilization which makes it adapt very well to the hypoxic plateau environment. Neuronal nitric oxide synthase (nNOS),one of three isoforms of nitric oxide synthase,plays an important role in the adapation to hypoxia. In
this study,the coding sequence of nNOS was cloned by RT-PCR from the brain of plateau zokor. The size of this gene is 4 290 bp in length,and encodes a protein composed of 1 429 aa. The nNOS gene of the plateau zokor was 87% to 90% homologous to human,dog,rabbit,mouse and rat. We identified typical nNOS domains,such as PDZ, oxidase, reductase domain and CaM binding sites within nNOSs of Plateau zokor. The molecular phylogenetic trees based on ML and Bayesian method showed that the plateau zokor clustered with mouse and rat,which was consistent with traditional phylogenetic relationship inferred from morphological and molecular markers. Three positive selected sites of Plateau zokor (332 T,1200 G and 1334 P)were identified by the PAML analysis,although statistically insignificant. Our results lay a foundation for further exploring the expression pattern,function and regulatory mechanism of nNOS in Plateau zokor acclimatization to hypoxia.

Key words: Gene cloning, Neuronal nitric oxide synthase, Plateau zokor (Myospalax baileyi), Sequence analysis

摘要: 高原鼢鼠是青藏高原特有的地下鼠,受到高原低氧以及洞穴低氧的双重低氧环境压力。经RNA 提取、RT-PCR、亚克隆与测序,本研究获得高原鼢鼠神经型一氧化氮合酶(nNOS)的编码区序列,并对其分子特征进行了分析。结果显示:高原鼢鼠nNOS 基因编码区(CDS)全长4 290 bp,编码1 429 个氨基酸残基;CDS 与大鼠、小鼠、兔、狗、人的同源性分别为90% 、89% 、87% 、87% 、89% ;结构域上,高原鼢鼠nNOS 具有PDZ蛋白结构域、氧化域、还原域及钙调素结合位点等nNOSs 所具有的典型结构域;基于nNOS 的最大似然树和贝叶斯树均支持高原鼢鼠与大鼠、小鼠具有最近的亲缘关系,与形态或其它分子标记构建的进化关系相符;分子进化分析检测到高原鼢鼠nNOS 中存在3 个正选择位点———332 T、1200 G 和1334 P,但均未达到统计显著水平。本研究为揭示高原鼢鼠nNOS 的表达特征及其在低氧适应中的作用与调控机制研究奠定了初步基础。

关键词: 高原鼢鼠, 神经型一氧化氮合酶, 基因克隆, 序列分析