【作者】 郑新欣；

【导师】 胡英考；

【作者基本信息】 首都师范大学 ， 遗传学， 2009， 硕士

【摘要】 为了获得具有组织表达特异性和与胁迫相关的小麦组蛋白H3,进一步了解组蛋白H3的功能。本文以拟南芥组蛋白H3的蛋白序列为信息探针,对小麦(Triticum aestivum)EST数据库进行同源检索筛选,克隆了小麦组蛋白H3基因。通过生物信息学的方法对得到的小麦组蛋白H3基因进行分析,分析的内容包括:内含子形式,电子基因表达谱和电子染色体定位。并采用半定量RT-PCR的方法分析了小麦组蛋白H3基因在盐处理下的表达情况。研究结果如下:1小麦组蛋白H3基因的克隆以拟南芥组蛋白H3的蛋白序列为信息探针,BLAST检索GenBank中的小麦EST数据库,获得了小麦组蛋白H3基因的cDNA序列。为了进一步验证电子克隆序列的正确性,在起始密码子和终止密码子区域设计特异引物,经RT-PCR进行克隆、序列分析验证,最终得到7个小麦组蛋白H3基因且与电子克隆序列相一致。经过分析,这7个基因所编码的小麦组蛋白H3可以分为两类:H3.1型和H3.2型。2小麦组蛋白H3基因的生物信息学分析以普通小麦基因组DNA为模板,扩增上述7个基因基因组序列,获得了3个基因的基因组序列,有一个基因含有一个长度为198bp的内含子,该内含子较其它物种类似基因内含子长一些。采用UNIGENE数据库进行的组织表达谱分析表明,克隆的基因组织特异性表达不明显。采用GrainGenes数据库将这7个基因定位于7BL,4AL和7AS染色体上。3小麦组蛋白H3基因在盐胁迫下表达量分析采用半定量RT-PCR的方法,对所获小麦组蛋白H3基因在盐胁迫处理下的表达情况进行了分析,获得了一个随盐处理时间的延长表达量增高的组蛋白H3基因。更多还原

【Abstract】 In order to further undersranding the functions of histon H3,we hoped to abtain tissue specific expression or stress related histon H3 genes.In this paper,we used the EST Blast method to clone Triticum aestivum histon H3 gene,analyzed its expression changes under salt stress using semi-quatitative RT-PCR,and demonstrated its intron pattern,expression profile and in silico mapping through bioimformatic analysis.1 Cloning and analysis of Triticum aestivum histon H3 geneSeven Triticum aestivum histon H3 gene was cloned by Blasting the Triticum aestivum EST database with the Arabidopsis thaliana homologous protein histom H3 sequence as a querying probe.To examine the accuracy of in silico cloning cDNA sequences,these sequences were cloned by RT-PCR and sequenced.The result confirmed that the in silico cloning cDNA sequences were accurate,and they can derided into two kind:type H3.1 and type H3.22 in silico analysis of Triticum aestivum histon H3 geneIn order to deternine the intron pattern of wheat histon H3 gene,specific primers which were designed for cDNA cloning were also used to amplify genomic DNA, three out of seven genomic sequences of histone H3 genes were abtained,and only one of them had one 198 bp intron,which is a little longer than those homolog genes in other species.In silico expression patterns in different tissues of those seven wheat histon H3 genes were investigated by Unigene database,the results shows that they have no tissue specific expression.Also Graingene database was used to determine their In silico mapping,which shows they were distributed over chromesome 7BL, 4AL and 7AS.3 expression of Triticum aestivum histon H3 gene in response to salt treatmentSemi-quantitative RT-PCR assay was performed to detect the expression pattern of the obtained Triticum aestivum histon H3 gene under salt stress,the result showed that the transcription of one histon H3 gene is accumulated after treated by salt.更多还原