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Title: Cloning and characterisation of a gene encoding sex-specific transcripts in Drosophila melanogaster
Author: MacDougall, Colin N.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1996
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In an attempt to identify genes involved in differentiation of sex-specific characteristics, a non-gonadal differential screen was performed. Sex-specific, radiolabelled cDNA was used to screen a bacteriophage λ genomic DNA library and a genomic recombinant, fs(1), was selected for further analysis on the basis of preferential hybridisation to female non-gonadal cDNA. in situ hybridisation to Drosophila 3rd instar larva salivary gland polytene chromosomes indicates that the genomic DNA contained within fs(I) is located distally, on the left arm of chromosome 3, at position 061C1-3. Two cDNAs, a 4.5kb cDNA (cDNA11) and a 3.0kb cDNA (cDNAa), were isolated on the basis of hybridisation to sequences contained within fs(1). Both cDNAs were fully sequenced and found to encode a novel OPA-repeat-containing serine/threonine-specific protein kinase. cDNAa and cDNA11 both contain the entire open reading frame (ORF) which encodes this predicted protein, and differ only in untranslated regions. The cDNAa ORF was subcloned into a fusion-protein expression vector and fusion protein was successfully expressed in bacterial cells, as shown by Western blot analysis using antibodies specific to the vector-derived fusion protein. Genomic DNA, containing the entire cDNA11 and cDNAa transcription units, was isolated. The precise intron/exon structure of both cDNAs was determined by Southern blotting, DNA sequencing and PCR analysis. cDNAa hybridises with four transcripts on Northern blots; a 3.0kb testis-specific, 3.5kb ovary-specific, 4.5kb female carcass-specific and a common transcript of around 4.7kb. The cDNA11-specific 3' UTR hybridises with the 4.5kb and common transcripts, but not with the 3.5kb or 3.0kb transcripts. Thus, cDNAa and cDNA11 are likely to represent the testis-specific and female carcass-specific transcripts, respectively. Both cDNAs contain translational control elements which are found in transcripts under male germline-specific translational control. The presence of these elements suggests that the testis-specific transcript is in fact germline-specific.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available