暗纹东方鲀中baff基因相关性序列的性质探讨生物化学与分子生物学专业论文.docxVIP

摘要摘要 摘要 摘要 本研究为解决暗纹东方啬屯养殖过程中病虫害的防治方法，从提高鱼体自身 免疫出发，探讨存在哪些与哺乳类、鸟类BlyS基因相关性的DNA序列以及这 些序列的性质。人B淋巴细胞刺激因子(又称为BAFF、BIyS、TALL．1、 zTNF4、THANK)是1999年发现的肿瘤坏死因子超家族的成员之一，它与某 些免疫疾病的致病机制密切相关，低量或过量表达均能引起机体的免疫失衡。 根据四种已知高度保守的BlyS同源片段设计引物，同时以草鱼、青鱼模板做外 对照，人模板做阳性对照，经PCR扩增、割胶回收、克隆测序。结果表明：以 鱼基因组为模板经PCR扩增获得了542bp、349bp、221bp三条片段；以人基因 组为模板获得5条片段，将最亮一条测序，发现是位于14号染色体上的片段， 与入BIyS没有相关性。542bp序列在暗纹东方纯和草鱼中是非常保守的，A含 量最低占18．8％，判断此段序列属于红鳍东方纯基因组文库中“N(任意核苷 酸)区，填补了红鳍东方纯基因组文库(2007)数据的空白。349bp和221bp在 暗纹东方啬屯、红鳍东方鲍及草鱼中都是保守的。因此542bp、349bp、221bp三 条片段属于引物特异性扩增，但对于BIyS来说是非特异性片段。暗纹东方纯和 红鳍东方纯序列比对分析显示：349bp序列中，转换颠换各发生3次、1次，插 入／缺失12次。221bp序列中，碱基替换各1次，插)k／缺失共6次，以上结果符 合大多数生物DNA序列进化中转换大于颠换的现象，但插入或缺失出现的频 率要远远高于碱基转换和颠换。在暗纹东方鲍和红鳍东方纯中碱基突变的主要 方式为插入或缺失。同时用生物学软件将此三段DNA序列按照三种读码框自 动翻译成氨基酸序列，能够折叠成类似BlyS蛋白空间结构的可能性很小，在蛋 白质数据库中做Blast，同样没有发现与BIyS蛋白相关性序列。结论：此三条 序列与BIyS蛋白的相关性很小，并且也不属于其他性质的基因或蛋白，推测可 能是新发现的三段序列，有待进一步研究。 关键词：暗纹东方纯；B淋巴细胞刺激因子；分子进化；红鳍东方纯 AbstractAbstract Abstract Abstract The research aimed to explore whether there was BlyS gene in the genome of Fugu obscurus，which exists in mammals and birds．Human B lymphocyte stimulator(also known as BAFF，BlyS，TALL-1，zTNF4，THANK)WaS found and considered to be one member of tumor necrosis factor(TNF)family．It Was closely related to pathogenic mechanisms of immue system diseases．Low level or excessive expression of it all call induce immune imbalance,Specific primers were designed based on the reported conserved BIyS gene regions，which were from four species：Homo sapiens， Mus musculus，Sus scrofa and Gallus gallus．Ctenopharyngodon idellus and Mylopharyngodon piceus were taked舔external control group．Homo sapiens were positive contr01．By PCR amplification,purifycation and withdraw,the products were cloned into pUCm·T vector and transformed into E．coli and sequenced．Three specific bands were obtained through PCR amplification,with the size 542，349 and 22 1 bp． 542bp—sequence was highly conserved in Fugu obscurus and Ctenopharyngodon idellus．The A content of 542bp-sequence Was 1 8．8％and the low