【作者】 杨龙；

【导师】 刘娣；

【作者基本信息】 东北农业大学 ， 动物遗传育种与繁殖， 2008， 硕士

【摘要】 MEF2家族属于MADS超家族,广泛的存在于骨骼肌与心肌细胞中,是最早发现具有肌肉调控特性的DNA结合活性因子。MEF2是肌肉组织发生和发育过程中的重要调控因子,控制脊椎动物肌肉特异基因表达,在肌纤维发育和神经系统发育中都起重要调节作用。基于肌细胞增强子在调控细胞增殖和肌肉分化中的重要作用,进一步揭示其功能和作用机理,有助于更好的理解肌肉组织的发生与发育模式。本研究采用比较基因组学和生物信息学等方法,克隆MEF2A基因并对其结构进行分析;通过构建分子进化树来研究MEF2A基因的系统发育;采用Real time PCR方法,研究了纯种大白猪MEF2A基因不同组织不同发育阶段的表达规律和MEF2家族4个基因的组织表达规律。主要研究结果如下:1.克隆了猪MEF2A基因cDNA序列(Genbank Accession Number:EF576923),基因编码区全长1524 bp,编码508个氨基酸,分子量为50844.98 Da,其结构上存在一个MADS-box的结构域、位于10-243附近。2.对猪MEF2家族基因的4个基因进行多重序列比对,发现一个极高的同源性区域,此处主要为MADS-box区。4个基因的MADS保守结构域同源比对分析,表明MEF2B与MEF2D的MADS-box结构域的同源性最高为100%,MEF2A与MEF2B和MEF2D的MADS-box结构域分化程度最大,同源性为89%。3.基于猪MEF2A基因的cDNA序列,依据Neighbor-Joining算法,构建了9个物种分子进化关系树。4.采用Real time PCR的方法对90、180、270日龄纯种大白猪的MEF2A基因心、肝、胃、脾、肾、肺、大肠、小肠、背肌、腿肌10个组织表达量进行了检测:表明MEF2A基因在肌肉组织中表达量较高且稳定,在其他组织中表达量较低,肝中的表达量最低。5.同样方法对90日龄MEF2家族4个基因进行了10个组织的表达量检测,4个基因组织表达上具有共同特征:肌肉组织(背肌、腿肌、心肌)表达量较高,脏腑肌(胃、大肠、小肠)表达量次之,其他组织中表达量较低;同时,也对同一组织中4个基因的表达情况进行了比较。更多还原

【Abstract】 MEF2 gene family belongs to the family of MADS (MCM1, agamous, deficiens, serum response factor)-box transcription factors, MEF2 exist in a wide range of cardiac and skeletal muscle cells, is the first discovered DNA binding activity factor with the muscle property, is an important regulatory factor for muscle histogeny and development. It can control vertebrate muscle-specific gene expression, and it is important for the process of muscle fiber and nervous system development. Because of the MEF2 plays an essential roles in regulating cell multiplication and tissue differentiation, to further reveal the function and mechanism of MEF2, contribute to better understanding the occurrence and development mode of muscle tissue. In this study, cloning the MEF2A and analysis its structure by comparative genomics and bioinformatics methods, phylogenesis of MEF2A also be analyzed by drawing Phylogentic tree; Expression of MEF2A gene and MEF2 gene family in different tissues and diferent period on large white pigs were analyzed by Real time PCR; The main results are as follows:1. MEF2A gene of pig was cloned (Genbank Accession Number: EF576923). The coding region of pig MEF2A gene is composed of 1524bp encoding 508 AA, and molecular weight is 50844.98 Da. There is a MADS-box domain in the vicinity of 10-243.2. There find a MADS-box domain by comparison of homology in the pig MEF2 gene family. The comparison of homology on MADS-box domain in four genes show that: the homology of MEF2B and MEF2D is 100%; MEF2A has large differentiation level, the homology with MEF2B and MEF2D is 89%.3. Based on the cDNA sequence of pig MEF2A gene, phylogenesis of nine species MEF2A be analyzed by drawing Phylogentic tree with Neighbor-Joining method.4. Expression of MEF2A gene in 10 tissues (Heart, Liver, Stomach, Spleen, Kidney, Lung, Large Intestine, Small Intestine, Dorsal muscle, Lower Limb muscle ) on 90, 180, 270-day-old large white pigs were analyzed by Real time PCR. The results show that the expression of MEF2A gene in muscle tissue is high and stable, and other organizations is relatively low, Liver is lowest.5. Expression of MEF2 gene family in 10 tissues on 90-day-old large white pigs were analyzed by the same mothed. The results showed that expression of four genes with common features: There is high expression in muscle tissue, eg: Dorsal muscle, Lower Limb muscle and Heart muscle; the second is visceral muscle, eg: Stomach, Large Intestine and Small Intestine; low expression in Liver, Kidney and Lung。At same time,compared the expression of MEF2 gene family in same tissue.更多还原