【作者】 王春生； 刘欣； 梁洋； 安铁洙；

【Author】 WANG Chun-sheng LIU Xin LIANG Yang AN Tiezhu College of Life Science,Northeast Forstry University,harbin 150040,China

【机构】 东北林业大学生命科学学院；

【摘要】 根据GenBank上蜘蛛拖丝蛋白基因序列(AY55585和AH015065)、拖丝蛋白基因的结构特点和密码子的简并性,设计378bp的拖丝蛋白基因单体,并对其人工聚合成四聚体.Primer5.0分析结果表明,决定拖丝弹性和抗拉力的氨基酸(Gla和Ala)含量(41.43和18.22)接近天然丝蛋白氨基酸含量(42.81和26.32):利用Antheprot软件对四聚体编码氨基酸的二级结构预测结果显示,与丝蛋白的氨基酸链相近,由4个相同的部分排列而成,即β-片层(占48%)、12个α-螺旋间隔(占15%)、散在的转角(占12%)和若干不规则卷曲(占25%)。将上述4聚体与改造的pcDNA3.1载体(K2.10启动子)连接构建真核表达载体,并通过PCR和酶切鉴定。上述结果为进一步开展有关转蜘蛛拖丝蛋白基因在绵羊被毛中表达的研究奠定了基础。更多还原

【Abstract】 According to spider dragline silk protein gene sequence in GenBank(AY55585 and AH015065),structure characteristic of the gene and degeneracy of codon,378bp sequence was designed,artificially synthesized and tetra-polymerized,amount of Gla and Ala that decided elasticity and strength of synthesized gene had an analogy with that of natural dragline silk protein gene by Primer5.0 analyzing,second structure the amino acid sequence was forecasted by Antheprot software,results showed that the amino acid sequence was composed of the same four segments,includingβ-sheet(48%),α-helix(15%),dispersedβ-turn(12%) and several anomalistic coi1(25%),these structures accorded with structure characteristic of silk protein.Eukaryotic expression vector was constructed by linking the artificial synthetic spider dragline silk protein gene to hair follicle-specific promoter,PCR and enzyme digestion identification indicated the recombination plasmid was right.Above results were necessary to further research spider dragline silk protein gene expression in eukaryote.更多还原