CHROMOSOME STRUCTURE DURING DROSOPHILA DEVELOPMENT

描述：此应用程序建议继续并扩展 他对异染色质和欧盟-异染色质边界的研究 最初是艾伦·斯普拉德林实验室的博士后研究员。这个 研究人员正在使用开发的DP(1；f)1187微型染色体系统 斯普拉丁实验室在过去的几年里。这个1.3Mb的微染色体 显示了典型高等真核生物的所有特征 染色体，但它提供了不同寻常的实验机会。在工作中 作为博士后研究员，PI发现DNA序列在 等色色-异色色边界具有不同寻常的特征，在此 他描述了他的计划，以进一步描述这些 序列和异染色质。 该提案包含三个具体目标。在第一个中，PI将 继续他对异色限制性片段的表征 Dp1187在许多不同的组织中的表达。在以前的实验中， PI发现，例如，唾液中的 腺体和卵巢。PI将进行测绘实验，以进一步 描述这些染色体的结构和限制 碎片。在卵巢DNA的实验中，他将专注于了解 Dp1187片段在Southern上表达不足的两个原因 斑点。PI发现这些碎片不会随着 典型的效率以及有缩短的Dp1187分子 在这个样本中。PI还将检查唾液腺Dp1187。在……里面 该组织存在Dp1187序列的真实表达下 不是因为转会的问题。这些分子的性质 产生的下表示将以2D为特征 杂交分析。私人投资公司还建议检查Dp1187在 二倍体细胞(在胚胎和成像盘/脑中)。圆周率已经 有证据表明在胚胎DNA中Dp1187序列没有得到充分的表达 转移不良导致的Southern杂交和Dp1187片段 在胚胎DNA中发现了尺寸增大的现象。他一直在发展 从单倍体精子中提取DNA的技术，他将 同时检查这些细胞中Dp1187的结构。 第二个具体目标是检查以下结果的一般性 Dp1187通过检查其他EU-异染色质边界。其中之一 他将检查的是作为亲本的sc8倒位。 Dp1187的染色体。在其他实验中，他将检查欧盟- 由于插入P元素而产生的异色边界 变成常染色体异染色质。 第三个具体目标是识别和表征物理 Dp1187异染色质的选择性转移特性 抑制力。他已经在溶液中分离出了耐转移DNA 他将以多种方式处理这种DNA，例如碱性 变性，然后以各种方式检查效果，例如 通过检查它在蔗糖梯度中的沉积来寻找改变 构象。

DESCRIPTION: This application proposes to continue and extend the research on heterochromatin and the eu-heterochromatin boundary that he started as a Postdoctoral Fellow in Allan Spradling's laboratory. The investigator is using the Dp(1;f)1187 minichromosome system developed in the Spradling lab over the last few years. This 1.3 Mb minichromosome displays all of the characteristics of a typical higher eukaryotic chromosome, but it affords unusual experimental opportunities. In work done as a postdoctoral fellow the PI found that DNA sequences at the euchromatic-heterochromatic boundary have unusual features and in this application he describes his plans to further characterize these sequences and heterochromatin in general. The proposal contains three specific aims. In the first the PI will continue his characterization of heterochromatic restriction fragments of Dp1187 in a number of different tissues. In previous experiments the PI found that there were differences between, for example, salivary gland and ovary. The PI will carry out mapping experiments to further characterize the structure of these chromosomes and restriction fragments. In the experiments on ovary DNA he will focus on understanding the two reasons that Dp1187 fragments are under represented on Southern blots. The PI has found that these fragments do not transfer with typical efficiency and as well that there are shortened Dp1187 molecules in this sample. The PI will also examine salivary gland Dp1187. In this tissue there is a true under representation of Dp1187 sequences that is it is not due to transfer problems. The nature of the molecules that give rise to the under representation will be characterized by 2D hybridization analysis. The PI also proposes to examine Dp1187 in diploid cells (in embryos and in imaginal discs/brains). The PI already has evidence that in embryo DNA Dp1187 sequences are under represented on Southern blots due to poor transfer and that Dp1187 fragments of increased size are found in embryo DNA. He has been developing techniques for the isolation of DNA from haploid sperm and he will examine the structure of Dp1187 in these cells as well. The second specific aim is to examine the generality of the results on Dp1187 by examining other eu-heterochromatin boundaries. One of those that he will examine is the sc8 inversion which is the parental chromosome for Dp1187. In other experiments he will examine eu- heterochromatic boundaries that result from the insertion of a P element into autosomal heterochromatin. The third specific aim is to identify and characterize the physical property of Dp1187 heterochromatin that leads to selective transfer inhibition. He has already isolated transfer resistant DNA in solution and he will treat this DNA in a variety of ways, such as alkaline denaturation and then examine the effects in a variety of ways such as by examining its sedimentation in sucrose gradients to look for altered conformation.