CONTROL OF TRANSCRIPTION OF ISOLATED EUCARYOTIC GENES

拟议研究的目标是确定地点和 目前正在调查这一问题的各个方面。基因调控 酵母甘油醛-e-磷酸脱氢酶(GAPDH)和烯醇酶基因。 研究了两种细菌的转录调控机制。 基于三个GAPDH基因和两个烯醇酶基因的一级结构 转录起始过程中程序化模式的控制 使用在体外构建的含有编码区缺失的质粒 肌动蛋白基因在酿酒酵母中的表达和功能调控。 转化和基因替换。含有缺失的突变株 这些反应统称为RNA加工。现在很清楚的是，RNA 已经使用替换方法进行了分离。突变株含有 最近对真核基因结构的阐明揭示了一种新的 分析每个基因重复序列的表达情况。GAPDH和GAPDH的表达 非编码序列称为插入序列或内含子。这些 存在于基因5‘和3’非编码区的序列决定了 这被称为RNA剪接。我们是第一个演示RNA拼接的人 含有GAPDH基因5‘非编码序列的基因融合 继续研究tRNA剪接反应并提出程序 5‘非编码序列是决定基因表达效率的关键因素 最近开发出允许在体外改变基因和它们的 将在体外构建烯醇化酶和GAPDH基因的非编码序列 诱变策略要求通过以下方式有机合成寡核苷酸 突变基因在酵母基因组中的适当位置的整合。 利用这些技术合成寡核苷酸以生产精确的 基因的非编码区。微生物的快速定量检测 综合。 基因已经被开发出来。最后，提出了序列需要 对于依赖于RNA聚合酶I的位点特异性转录启动 体外靶向鉴定克隆的酵母核糖体基因 诱变作用及RNA选择性转录所需成分 聚合酶I被分离了。这次调查的长期目标是 会在体外重建酵母的选择性转录 糖酵解和核糖体基因。

The goals of the proposed research are to determine the location and aspects of this problem are currently being investigated. Control of gene yeast glyceraldehyde-e-phosphate dehydrogenase (GAPDH) and enolase genes. have studied the mechanisms of transcriptional control in tow bacterial be based on the primary structures of the three GAPDH and two enolase genes the control of the programmed pattern of transcriptional initiation during plasmids containing coding region deletions, constructed in vitro, be used control of expression and the function of the actin gene in S. cerevisiae. transformation and gene replacement. A mutant strain containing a deletion reactions collectively known as RNA processing. It is now clear that RNA already been isolated using the replacement method. Mutants containing recent elucidation of eukaryotic gene structures has revealed a new analyze expression of each gene repeat. The expression of GAPDH and non-coding sequences known as intervening sequences or introns. These sequences present in the 5' and 3' noncoding regions of the genes determine called RNA splicing. We are the first to demonstrate the RNA splicing gene fusion containing the 5' noncoding sequences of a GAPDH gene and the continuing the study of the tRNA splicing reaction and propose procedures 5' noncoding sequences are responsible for the efficiency of expression of recently been developed allowing the in vitro alteration of genes and their noncoding sequences of enolase and GAPDH genes will be constructed in vitro mutagenic strategies requires the organic synthesis of oligonucleotides by integration of the mutant gene at its proper locus in the yeast genome. to use these techniques to synthesize oligonucleotides to produce precise noncoding regions of the gene. Rapid quantitative assays for the synthesis. gene have been developed. Finally, it is proposed that sequences required for RNA polymerase I dependent site specific initiation of transcription in vitro on cloned yeast ribosomal genes be identified by targeted in vitro mutagenesis and that components required for selective transcription by RNA polymerase I be fractionated. The long term goal of this investigation will be in vitro reconstitution of selective transcription of yeast glycolytic and ribosomal genes.