EXPRESSION OF GENES INJECTED INTO XENOPUS LAEVIS

我们打算研究两个因素涉及的监管， 真核基因的转录。 其中包括：（1）功能 位于基因5'端的序列和（2）基因在 染色体 最近的证据表明， 与许多真核基因编码区的5 ′端相邻（即， CCAAT，TATA）影响转录的速率和特异性。 入会仪式 其他的结果表明， CCAAT和TATA也影响或调节个体的表达， 基因（即，海胆组蛋白基因）。 我们将检验这个假设， 调节序列位于距离5'端几百个核苷酸的 果蝇醇脱氢酶基因调节这种 基因 我们将在5'端创建一系列体外缺失突变， 果蝇ADH基因的限制性内切酶， 分子克隆 对这些特异序列的功能进行分析 通过将修饰的基因显微注射到非洲爪蟾卵母细胞的细胞核中， 随后进行转录物的5 ′和3 ′ S1核酸酶作图。 先前 研究已经分析了编码普遍存在的功能的基因。 结果 这项研究将使我们能够确定调制器序列是否存在于一个 编码分化功能的基因，并鉴定该序列。 为了制定基本原则，这些信息是必要的 基因调控。 我们有证据表明DNA序列 显微注射到受精卵中的基因被整合到非洲爪蟾的基因组中。 本提案的第二部分将决定网站的效果 海胆组蛋白基因和果蝇ADH基因的整合 其发育和组织特异性表达。 这将是 通过显微注射果蝇ADH和海胆组蛋白基因 非洲爪蟾的受精卵，并分析发育和 当这些基因整合到不同的组织中时， 在基因组中的位置。 我们觉得这个制度特别好， 适合这些研究，因为我们有证据表明， 序列，并设计了一个系统，我们可以生产克隆或 青蛙通过核移植，其中每个克隆将包含 注入的基因整合在不同的染色体位点。 的结果 这项研究将对基因的性质产生深远的影响。 监管机制以及与健康有关的遗传程序。

We intend to examine two factors involved in the regulation of transcription of eukaryotic genes. These include: (1) the function of sequences located 5' to a gene and (2) the physical location of the gene in the chromosome. Recent evidence has shown that sequences immediately adjacent to the 5' end of the coding region of many eukaryotic genes (ie. CCAAT, TATA) affect the rate and specificity of transcriptional initiation. Other results suggest that sequences further upstream from the CCAAT and TATA also influence or modulate the expression of individual genes (ie. sea urchin histone genes). We will test the hypothesis that modulating sequences located several hundred nucleotides from the 5' end of the Drosophila Alcohol dehydrogenase gene modulate the expression of this gene. We will create a series of in vitro deletion mutations at the 5' end of the Drosophila ADH gene utilizing restriction endonucleases and molecular cloning. The function of the specific sequences will be analyzed by microinjection of the modified gene into the nuclei of Xenopus oocytes followed by 5' and 3' S1 nuclease mapping of the transcripts. Previous studies have analyzed genes coding for ubiquitous functions. The results of this study will enable us to determine if modulator sequences exist in a gene coding for a differentiated function and to identify the sequence. This information will be necessary in order to formulate basic principles of gene regulation. We have evidence which suggests that DNA sequences microinjected into fertilized eggs are integrated into the Xenopus genome. The second portion of this proposal will determine the effect of the site of integration of both sea urchin histone genes and the Drosophila ADH gene on their developmental and tissue specific expression. This will be approached by microinjecting Drosophila ADH and sea urchin histone genes into fertilized eggs of Xenopus laevis and analyzing the developmental and tissues specific expression of these genes when integrated into different positions in the genome. We feel that this system is especially well suited for these studies since we have evidence for integration of injected sequences, and have devised a system by which we can produce clones or frogs by nuclear transplantation in which each clone will contain the injected gene integrated in a different chromosomal site. The results of this study will have far reaching implication regarding the nature of gene regulatory mechanisms as well as health related genetic procedures.