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Title: Cloning and characterization of the ubiquitin carboxyl terminal hydrolase gene in Drosophila melanogaster
Author: Zhang, Nian
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1992
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In this project, 61 strains of Drosophila melanogaster carrying the enhancer detector P[lac, ry+]A were stained for the β-galactosidase activity in adult gonad. This P-transposon detects regulatory elements of a nearby gene. Sex- and tissue-specific staining were found in many strains. Some strains showed staining in very specific cells. These cells may play very important roles during oogenesis. 16.5 kb genomic fragment surrounding the enhancer detector P[lac, ry^+], located at 98F of chromosome 3, has been cloned from a strain called D19. This strain shows the lacZ staining in germ-line cells of adult ovary and testis. This DNA fragment contains multiple transcription units. A 1.3 kb HindIII subfragement GH4 contains a gene encoding a 1.1 kb mRNA. A corresponding cDNA ovD19C2 has been isolated from a Drosophila ovary cDNA library. Northern analysis shows that this gene is expressed at low levels during most development stages, while the transcription is enhanced in the ovary and testis. High level mRNA of this gene is also presented in the early embryos. In situ hybridization shows that the enhanced transcription occurs in nurse cells and spermatocyte cysts. The high level mRNA of this gene in the early embryo must be synthesised in the nurse cells and transported and stored in the oocyte. Sequencing analysis shows this gene contains three introns. The longest open reading frame of the cDNA encodes a protein of 227 amino acid residues. The amino acid sequence of the predicted protein is highly identical to the human ubiquitin carboxyl terminal hydrolases. It has 51.1% identity to the human ubiquitin carboxyl terminal hydrolase isozyme L3 (UCH-L3), and 47% identity to isozyme L1 of the same enzyme. The predicted protein of ovD19C2 also has many important features which are common in this enzyme family. Attempts were made to mutate this gene by imprecise excision of the P-transposon from the strain D19. 45 lines which are homozygous lethal for chromosome 3 have been obtained. Further characterization of these lines will provide very important information about the gene cloned.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available