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果蝇鸟氨酸脱羧酶基因座的分离与鉴定:果蝇基因组中存在两个转录的ODC基因的证据。

Isolation and characterization of the Drosophila ornithine decarboxylase locus: evidence for the presence of two transcribed ODC genes in the Drosophila genome.

作者信息

Rom E, Kahana C

机构信息

Department of Molecular Genetics and Virology, Weizmann Institute of Science, Rehovot, Israel.

出版信息

DNA Cell Biol. 1993 Jul-Aug;12(6):499-508. doi: 10.1089/dna.1993.12.499.

Abstract

The polymerase chain reaction (PCR) was used to isolate two Drosophila ornithine decarboxylase (ODC) genes. Two mixtures of degenerate oligonucleotides corresponding to peptides that are fully conserved among ODCs from widely diverged species were used as opposing primers in the PCR with cDNA or genomic DNA as templates. Sequence analysis of the resulting DNA products confirmed their identity as ODC fragments. The genomic PCR product was then used as a probe for screening a Drosophila genomic library, resulting in the isolation of genomic clones representing two distinct ODC genes (dODC1 and dODC2). Sequence analysis of both genes demonstrated that although varying at their coding and noncoding regions, their overall structure is extremely similar containing 6 exons and 5 short introns. Southern blot and sequence analyses revealed that the two ODC genes are arranged in a tandem head-to-tail configuration. Both ODC genes were assigned by in situ hybridization analysis to position 44A on the right arm of the second chromosome. The isolation of cDNA clones corresponding to these two ODC genes demonstrated that both are transcribed in the adult fly. We hope that the isolation of genomic and cDNA clones of Drosophila ODC will permit the investigation of the expression of ODC during Drosophila development and the role of polyamines in this process.

摘要

聚合酶链反应(PCR)被用于分离两个果蝇鸟氨酸脱羧酶(ODC)基因。两种与来自广泛不同物种的ODC中完全保守的肽段相对应的简并寡核苷酸混合物,被用作以cDNA或基因组DNA为模板的PCR中的反向引物。对所得DNA产物的序列分析证实它们是ODC片段。然后将基因组PCR产物用作筛选果蝇基因组文库的探针,从而分离出代表两个不同ODC基因(dODC1和dODC2)的基因组克隆。对这两个基因的序列分析表明,尽管它们在编码区和非编码区存在差异,但其总体结构极为相似,包含6个外显子和5个短内含子。Southern印迹和序列分析显示,这两个ODC基因以头对尾的串联形式排列。通过原位杂交分析将这两个ODC基因定位到第二条染色体右臂的44A位置。对应于这两个ODC基因的cDNA克隆的分离表明,它们在成年果蝇中均有转录。我们希望果蝇ODC基因组和cDNA克隆的分离将有助于研究果蝇发育过程中ODC的表达以及多胺在此过程中的作用。

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