THE MOLECULAR BASIS OF POSITION EFFECT VARIEGATION

位置效应变异的分子基础

基本信息

批准号：
3284629
负责人：
ROSS Joseph MACINTYRE
金额：
$ 13.89万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1985
资助国家：
美国
起止时间：
1985-01-01 至 1989-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3284629
关键词：
Drosophilidae acid phosphatase alleles chromosome aberrations complementary DNA cytogenetics densitometry endonuclease enzyme mechanism gene expression gene mutation genetic manipulation genetic recombination genetic transcription heterochromatin messenger RNA molecular cloning molecular genetics mutagen testing nucleic acid hybridization nucleic acid probes nucleic acid sequence structural genes

Drosophilidae acid phosphatase alleles chromosome aberrations complementary DNA cytogenetics densitometry endonuclease enzyme mechanism gene expression gene mutation genetic manipulation genetic recombination genetic transcription heterochromatin messenger RNA molecular cloning molecular genetics mutagen testing nucleic acid hybridization nucleic acid probes nucleic acid sequence structural genes

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项目摘要

The basis for the phenomenon of position-effect variegation in Drosophila melanogaster will be investigated using a cloned acid phosphatase-1 (Acph-1) gene as a molecular probe. The expression of the Acph-1 gene, when it has been relocated within or near broken heterochromatin, will be monitored at the level of mRNA production. In addition, the gene copy number in cells with polytene chromosomes will be measured to determine if reductions in mRNA are associated with lowered levels of genic DNA. These results will be compared to those obtained from cells with non-polytene chromosomes. The state of the chromatin adjacent to variegating and non-variegating Acph-1 genes will be analyzed with a series of probes which both surround and include the Acph-1 gene. This will be done in rearrangements which differ with regard to the distance between the breakpoint and the locus and/or the severity of the genic inactivation associated with the breakpoint. DNA spanning the heterochromatic-euchromatic junctions will be cloned, restriction mapped and sequenced in order to detect features common in those rearrangements which induce position effects. Finally, in transformation experiments the sequences and/or the amount of heterochromatin necessary to cause position effects will be determined using the Acph-1 gene as a model system. These experiments should help us understand the ways in which altered chromatin states affect gene activation as well as providing fundamental information about the causal basis of position effect variegation.

果蝇位置效应变化现象的基础将使用克隆的酸性磷酸酶1研究Melanogaster （ACPH-1）基因作为分子探针。 ACPH-1基因的表达，当它被重新定位在或骨折的异染色质中时，在mRNA产生水平上进行监测。另外，基因副本将测量带有多烯染色体的细胞中的数量，以确定是否是否 mRNA的降低与遗传DNA水平降低有关。这些结果将与从非聚丁烯细胞获得的结果进行比较染色体。染色质的状态与杂色和将使用一系列探针分析非变速箱ACPH-1基因既包围并包括ACPH-1基因。这将在重排在距离之间的距离断点和基因座和/或基因失活的严重程度与断点关联。 DNA跨越杂色 - 纤维通交连接将被克隆，限制映射并测序以检测这些重排中常见的特征诱导位置效应。最后，在转换实验中序列和/或引起位置所需的异染色质量效果将使用ACPH-1基因作为模型系统确定。这些实验应该帮助我们了解改变的方式染色质状态影响基因激活并提供基本有关位置效应变化的因果基础的信息。