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| 兰州大尾羊心脏型脂肪酸结合蛋白（H-FABP） 基因克隆及其同源性比较 | |
| 引用本文： | 徐红伟,柏家林,冯玉兰,曹忻,蔡勇,金方圆,达小强,杨具田,臧荣鑫.兰州大尾羊心脏型脂肪酸结合蛋白（H-FABP） 基因克隆及其同源性比较[J].中国农业科学,2013,46(3):639-646. |
| 作者姓名： | 徐红伟 柏家林 冯玉兰 曹忻 蔡勇 金方圆 达小强 杨具田 臧荣鑫 |
| 作者单位： | 1. 西北民族大学实验中心,兰州,730030 2. 西北民族大学生命科学与工程学院,兰州,730030 3. 西北民族大学实验中心,兰州730030;西北民族大学生命科学与工程学院,兰州730030 |
| 基金项目： | 国家自然科学基金（31160440，31260533）、甘肃省科技支撑计划项目（1011NKCA051）、甘肃省农业生物技术研究与应用开发项目（GNSW-2009-13）、国家民委科研项目（2009-158-09XB02）、兰州市科技计划项目（2011-1-113） |
| 摘 要： | 【目的】克隆兰州大尾羊心脏型肪酸结合蛋白（H-FABP）基因全长cDNA序列，为研究绵羊H-FABP生物学作用和生产应用提供理论依据。【方法】根据已知哺乳动物H-FABP基因 cDNA 序列，设计5''和3''特异引物，运用cDNA 末端快速扩增（RACE）技术获得兰州大尾羊H-FABP基因全长 cDNA 序列。【结果】 扩增获得兰州大尾羊5''端425 bp、3''端231 bp片段和 177 bp中间片段，拼接获得748 bp兰州大尾羊H-FABP基因全长cDNA 序列（GenBank登录号：JQ780322）。 兰州大尾羊H-FABP基因ORF长 402 bp，编码 133 个氨基酸。核苷酸序列分析显示兰州大尾羊H-FABP基因序列与大多数哺乳动物相似，但其第66位发生的碱基转换（T←→G）引起所编码的第22位天门冬氨酸（N）不同于其它所有物种的赖氨酸（K）。构建的基因进化树分析结果显示兰州大尾羊与山羊亲缘关系最近。预测兰州大尾羊H-FABP蛋白质的空间结构与山羊和牛H-FABP类似，由2个α螺旋和10个反向平行的β折叠组成，10 个折叠片围成一个桶状结构，疏水性残基位于桶内，用于结合脂肪酸。【结论】克隆了兰州大尾羊H-FABP基因，为进一步研究该基因的功能奠定了基础。 |
| 关 键 词： | 兰州大尾羊 H-FABP基因 cDNA 末端快速扩增 序列分析 |
| 收稿时间： | 2012-10-30 |
Cloning and Sequence Analysis of the Full-Length cDNA of H-FABP Gene in Lanzhou Fat-Tailed Sheep |
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| XU Hong-Wei,BAI Jia-Lin,FENG Yu-Lan,CAO Xin,CAI Yong,JIN Fang-Yuan,DA Xiao-Qiang,YANG Ju-Tian,ZANG Rong-Xin.Cloning and Sequence Analysis of the Full-Length cDNA of H-FABP Gene in Lanzhou Fat-Tailed Sheep[J].Scientia Agricultura Sinica,2013,46(3):639-646. | |
| Authors: | XU Hong-Wei BAI Jia-Lin FENG Yu-Lan CAO Xin CAI Yong JIN Fang-Yuan DA Xiao-Qiang YANG Ju-Tian ZANG Rong-Xin |
| Institution: | 1.Science Experimental Center, Northwest University for Nationalities, Lanzhou 730030; 2.College of Life Science and Engineering, Northwest University for Nationalities, Lanzhou 730030 |
| Abstract: | 【Objective】To clone the full length cDNA of heart fatty acid-binding protein (H-FABP) gene in Lanzhou fat-tailed sheep for providing a theoretical basis to study its biological function and application in sheep. 【Method】 The 5′- and 3′- gene specific primers were designed according to the alignment of known cDNA sequences of H-FABP from mammals. Technique of rapid amplification of cDNA ends (RACE) was employed to clone the full length cDNA of H-FABP gene in Lanzhou fat-tailed sheep. 【Results】 About 425 bp 5′-RACE cDNA and 231 bp 3′-RACE cDNA was obtained by 5′-RACE and 3′-RACE, respectively, using skeletal muscle RNA transcribed cDNA as template. Nest PCR was performed to clone 177 bp intermediate fragment. The full length cDNA of 748 bp H-FABP gene was spliced (GenBank Accession Number JQ780322). The open reading frame of sheep H-FABP gene is 402 bp in length, encoding a mature protein H-FABP of 133 amino acids and a resulting Mr=14 761. Phylogenetic analysis showed that H-FABP gene in Lanzhou fat-tailed sheep is more close to goat, Capra hircus. Alignment comparison indicated that nucleotide homology of H-FABP gene in sheep is more similar with mammals. However, the base transition from T to G in sixty-six of nucleotide sequence leading to the change from asparagines (N) to lysine (K) in twenty-second of amino acid sequence, which is different from other species. It is predicted that tertiary structure of H-FABP protein is very similar to H-FABP of C. hircus, having 2 α-helix, 10 antiparallel β-pleated sheets that form barrel. 【Conclusion】The full length cDNA of 748 bp H-FABP gene was first to be cloned by RACE. This finding may provide basic data for further studying the role of H-FABP gene in sheep. |
| Keywords: | Lanzhou fat-tailed sheep H-FABP gene rapid amplification of cDNA ends (RACE sequence analysis |
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