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  • 1
    ISSN: 1573-4927
    Keywords: Drosophila grimshawi ; DNA length variants ; Drosophila Stock Center ; molecular evolution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Two V3 vitellogenin clones isolated from genomic libraries ofDrosophila grimshawi (G1, Auwahi, Maui) were found to differ in length. Structural comparison of the two clones established that the length difference could be attributed to two insertions/deletions of about 200 bp each, both within the 3′ flanking sequences of the gene. The two length variants appeared to be polymorphic in the G1 laboratory strain, as demonstrated by analysis of genomic DNA isolated from single flies. The deleted variant sequence was traced by further analysis to two otherD. grimshawi strains (PK9 and S10G1) which originated from the island of Molokai. The existence of this morph in the Maui strain appears to have resulted from a laboratory stock contamination at the Drosophila Stock Center. In the course of a few generations of culture of this G1 strain at New York University, the deleted morph increased its frequency surprisingly rapidly, almost replacing the original morph, while at the Bowling Green Stock Center, the original morph still predominates. These frequency changes are most likely consequences of genetic drift due to bottlenecks in the maintenance and propagation of this stock.
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  • 2
    ISSN: 1573-5028
    Keywords: ABA element ; carrot embryogenesis ; seed-specific factors ; Lea gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Many late embryogenesis abundant (Lea) protein genes in plants are regulated by abscisic acid (ABA). The RNA level of a carrot gene, DC8, increases in response to ABA in developing seeds. However, DC8 cannot be induced by ABA in adult tissues. We used chimeric genes made of various DC8 promoter fragments fused to β-glucuronidase (GUS) to analyze the transcriptional regulation of DC8. DC8:GUS expression was measured in electroporated carrot protoplasts and in stably transformed carrots. The region of the DC8 promoter from −170 to −51 contained ABA-responsive sequences that required a 5′ upstream region for high levels of expression in embryogenic callus protoplasts. 505 bp of the DC8 promoter conferred GUS expression in stably transformed somatic and zygotic embryos. DC8:GUS was expressed only in tissues formed in the seed. This includes cells in the embryo, the endosperm and the germinating seedlings. Gel retardation and competition experiments were performed to analyze the embryo nuclear protein-DNA binding activities in vitro. No binding activity was detected on the putative ABA-responsive region; however the 5′ upstream regions located between -505 and -301 interacted with embryo nuclear factors. An additional site of DNA-protein interaction was located between positions -32 and +178. The nuclear proteins that bind these sequences were found in the embryo nuclei only, not in the nuclei from leaves or roots.
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  • 3
    ISSN: 1573-5028
    Keywords: abscisic acid ; ABA-response element ; bi-directional promoter ; Brassica napus ; oleosin ; seed development
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In Brassica napus, oleosins are expressed at high levels in the seed during the latter stages of embryo development. The cis-acting regulatory properties of an 872 bp promoter fragment of a B. napus oleosin gene were examined by analysis of β-glucuronidase (GUS) expression in transgenic tobacco plants containing an oleosin promoter-GUS transcriptional fusion. The reporter gene was expressed at high levels only in seeds, specifically in embryo and endosperm tissue and regulated throughout seed development. These data demonstrate that oleosin gene transcription is regulated in a tissue-specific and temporally regulated manner and clearly indicate that oleosin protein expression is co-ordinated primarily at the transcriptional level. Oleosin mRNA was shown to be abscisic acid (ABA) inducible and an ABA-response element in the oleosin promoter was shown to be bound by a protein factor in a sequence-specific manner. Sequence analysis of the oleosin promoter has identified several other putative cis-acting sequences which may direct oleosin gene expression. The presence of a large open reading frame in the bottom strand of the oleosin promoter (ORF2) which encodes a polypeptide similar to the ethylene-induced E4 gene of tomato is reported. A PCR-generated DNA probe containing the ORF2 sequence hybridised with a 1.4 kb transcript in total RNA extracts of a variety of tissues, including leaves and germinated seed cotyledons. This finding suggests that the oleosin gene promoter directs transcription in both directions. It is the first report of a bi-directional nuclear gene promoter in plants.
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  • 4
    ISSN: 1573-5028
    Keywords: anther ; Brassica ; β-glucuronidase ; oleosin ; promoter ; tapetum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract To investigate the sequences responsible for the regulated expression of tapetal-specific oleosin-like genes, ca. 2 kb of the 5′-upstream regions from two divergent genes, OlnB;4 and OlnB;13, were isolated, sequenced and fused to the reporter gene β-glucuronidase for study in transgenic Brassica napus plants. Although the proteins encoded by these two genes are highly divergent, except for the conserved oleosin-like domain, the first 250 bp of their 5′-upstream regions was 86% identical, including a region of 150 bp upstream from the TATA box. Analysis of 42 independent transformants by histochemical and fluorometric methods showed that both promoters directed tapetal-specific expression that peaked at the 4 mm flower bud stage.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 218 (1989), S. 143-151 
    ISSN: 1617-4623
    Keywords: Carrot ; Somatic and zygotic embryos ; DNA sequence ; Gold immunolocalization ; Allelic variability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary To understand the morphogenetic and physiological processes occurring during plant embryogenesis, we isolated cDNA clones homologous to genes preferentially expressed during somatic embryogenesis. One of these cDNA clones detected an embryo-specific mRNA species with a corresponding protein of 66 kDa. The expression pattern of the mRNA is similar between somatic and zygotic embryos of carrots. To characterize the gene encoding this mRNA, we isolated the corresponding genomic clones. Molecular analysis of the DNA from several haploid and diploid carrots showed that the mRNA was encoded by a single copy gene, named DC 8. DNA sequence analysis showed that the gene consisted of three exons and coded for a hydrophilic protein with a central region composed of 17 repeats. At the NH2-terminus no typical signal sequence was found. Immunocytochemical analysis localized the protein primarily in the vacuoles and protein bodies of zygotic embryos; the cytoplasm showed some antibody staining. The protein was also found in cell walls of endosperm tissue. The amount of DC 8 protein was too low for it to be categorized as a seed storage protein; its role in embryogenesis remains to be determined.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 206 (1987), S. 475-484 
    ISSN: 1617-4623
    Keywords: Drosophila grimshawi ; Vitellogenin genes ; Structure
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We isolated recombinant genomic DNA clones containing sequences coding for the female-specific vitellogenin proteins of Drosophila grimshawi. By screening with cDNA vitellogenin clones derived from female fat body mRNA we were able to isolate all three genes, namely V1, V2 and V3. The identity of these genes was established first by cell-free translation of the hybrid-selected mRNA followed by protease digestion of the in vitro translation products and second by hybridization of the three genes to electrophoretically separated mRNAs. The transcriptional orientation of these genes was determined. The V1 and V2 genes have opposite orientations with their 5′-ends 1.75 kb apart. S1 analysis demonstrated that the V1 gene has three exons of 310, 400 and 980 bp in length and two introns of about 120 bp. The V2 gene has two exons of 300 and 1260 bp in length and an intron 100 bp long. The V3 gene has three exons of 250, 375 and 820 bp in length and two introns of about 120 bp. The homology, in both sequence and structure, of the vitellogenin genes indicates that they have arisen by duplication events from an ancestral gene. Moreover, the similarity of the V1 and V2 gene positions within the genome of the two distant species D. melanogaster and D. grimshawi suggests a functional coupling of these two genes during vitellogenin gene expression.
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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 210 (1987), S. 564-571 
    ISSN: 1617-4623
    Keywords: Drosophila grimshawi ; Vitellogenin genes ; Expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The three vitellogenin transcripts from Drosophila grimshawi have an average size of 1,600 nucleotides as determined using denaturing electrophoretic conditions. Southern analysis showed that the large quantity of vitellogenin mRNAs in adult female fat body cells is not a reflection of specific gene amplification. The quantitative differences in mRNA accumulation between fat body and follicle cells, which are in concert with the onset of their translation, indicate that vitellogenin synthesis entails the regulated expression of individual genes. The expression of the vitellogenin genes during follicle development is stage specific: V1 and V2 expression starts at late stage 7, while V3 is delayed by one stage. Maximum transcription of all three genes occurs at stage 10 whereas at stage 12 none of the transcripts is present. These results suggest that, either there is more than one regulatory signal, or there is one to which each gene reacts differently. Surprisingly, in male fat body cells a V2 transcript has been detected which is also present in the poly(A)+RNA fraction: the function and the purpose of this particular vitellogenin mRNA in male fat body cells are unknown. Neither of the other two vitellogenin transcripts have been detected in male fat body cells.
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  • 8
    ISSN: 1617-4623
    Keywords: Transposable elements ; Genome organization ; Male recombination ; hobo ; GD sterility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary P strains of Drosophila melanogaster are characterized by the presence of both full-length and deletion derivatives of the transposable element P in their genome, and by their ability to induce the syndrome of hybrid dysgenesis among the progeny of certain intra-strain crosses, when introduced through the male parents. In contrast, strains belonging to the M' class, and which were also found to bear P element-homologous sequences, lack this ability and this has been attributed to the presence in the genome of most of these strains of a distinct class of deletion derivatives termed KP, which can suppress the action of functional P factors. Here we demonstrate that KP elements are present, next to full-length ones, in the genome of at least three strains which induce P-M-like dysgenic symptoms, including GD sterility. KP elements form the majority of the P-homologous sequences in the strains MR-h12, 23.5/CyL 4 and the latter's derivative 23.5 */Cy. While the first one is a genuine P strain and the second one depicts a strong P cytotype, the third is a genuine M' strain. The hybrid dysgenesis induced by the two 23.5 MRF strains seems to be due, not primarily to the P elements, but to the action of hobo elements.
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  • 9
    ISSN: 1573-5028
    Keywords: cytochrome b5 ; fruit and flower development ; gene expression ; in situ hybridisation ; olive
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report the characterisation of two cytochrome b5 genes and their spatial and temporal patterns of expression during development in olive, Olea europaea. A PCR-generated probe, based on a tobacco cytochrome b5 sequence, was used to isolate two full-length cDNA clones (cytochrome b5-15 and cytochrome b5-38) from a library derived from 13 WAF olive fruits. The cDNAs encoded proteins of 17.0 and 17.7 kDa, which contained all the characteristic motifs of cytochromes b5 from other organisms and exhibited 63% identity and 85% similarity with each other. The olive cytochrome b5-15 cDNA was then used as a probe for more detailed analysis. Southern blotting revealed a gene family of at least 4–6 members while northern blotting and in situ hybridisation showed a highly specific pattern of gene expression. Very low levels of cytochrome b5 mRNA were detected in tissues characterised by high rates of lipid accumulation, such as young expanding leaves, maturing seeds and ripening mesocarp. The cytochrome b5 genes were not induced at 6 °C and their response to ABA was relatively slow compared with fatty acid desaturase genes. In contrast, high levels of cytochrome b5 gene expression were found in young fruits at the pattern formation (globular/heart) stage of embryogenesis and in vascular and transmitting tissues of male and female reproductive organs. The data are consistent with a major role for cytochrome b5 in developmental processes related to plant reproduction in addition to being an electron donor to microsomal desaturases.
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 35 (1997), S. 955-961 
    ISSN: 1573-5028
    Keywords: Arabidopsis ; expression ; heat-shock ; hsp90 gene family ; sequence comparison
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have isolated six members of the hsp90 gene family from Arabidopsis thaliana. Three genes designated hsp81.2, 81.3 and 81.4 are clustered within a 15 kb genomic region while two of these are 1.5 kb apart in a head-to-head orientation. The deduced amino acid sequence shows that the members can be divided into two types. The hsp81.1, 81.2, 81.3 and 81.4 genes comprise the cytosolic hsp90 type having few introns. However, the hsp88.1 and 89.1 genes comprising the organelle type are composed of 18 or 19 introns. Sequence comparison showed there is high homology among the cytosolic members while there is less homology among the organelle members. The expression of the hsp90 genes and mRNA accumulation in plants and calli is very low at control temperatures and is strongly induced by heat-shock. Arsenite stress strongly stimulates the expression of this gene family.
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