RESUMEN

Odorant receptors (ORs) constitute the molecular basis for the detection of volatile odorous molecules and the perception of smell. Our understanding of chemical senses has been greatly expanded by the discovery of the OR gene families in vertebrates and in the nematode Caenorhabditis elegans. Recently, candidate Drosophila OR genes have been identified. The putative ORs do not possess any primary sequence identity with known vertebrate or C. elegans receptors, but belong to the family of G protein-coupled receptors according to their predicted seven transmembrane topology. To prove olfactory function of these proteins, we expressed a member of the putative Drosophila OR gene family, Or43a, in Xenopus laevis oocytes. Using two-electrode voltage-clamp recording we identified four odors (cyclohexanone, cyclohexanol, benzaldehyde, and benzyl alcohol) that activated the receptor at low micromolar concentration and structurally related substances that did not. This report shows the function and specificity of a member of the recently identified family of Drosophila ORs expressed in a heterologous system.

Asunto(s)

Drosophila/metabolismo , Receptores Odorantes/metabolismo , Animales , Secuencia de Bases , Cartilla de ADN , ADN Complementario , Femenino , Potenciales de la Membrana , Técnicas de Placa-Clamp , Receptores Odorantes/efectos de los fármacos , Receptores Odorantes/genética , Transducción de Señal , Xenopus laevis

RESUMEN

The hnRNP K protein is among the major hnRNA-binding proteins with a strong preference for cytidine-rich sequences. We have cloned a Drosophila hnRNP protein closely related to this vertebrate protein. The protein first identified by the monoclonal antibody Q18 is encoded by a gene located in 57A on polytene chromosomes and has been consequently named Hrb57A. The amino acid sequence of the Hrb57A KH domains and their overall organisation in the protein are remarkably similar to the vertebrate proteins. As the hnRNP K in vertebrates the M(r) 55 000 Drosophila Hrb57A/Q18 protein strongly binds to poly(C) in vitro and is ubiquitously present in nuclei active in transcription. On polytene chromosomes it is found in many puffs and minipuffs. Hrb57A/Q18 specifically coprecipitates four other proteins: Hrb87F/P11 a Drosophila hnRNP A1 homologue, the hnRNA-binding protein S5, the RNA recognition motif-containing protein NonA and the RNA-binding zinc finger-containing protein on ecdysone puffs PEP/X4.

Asunto(s)

Drosophila melanogaster/genética , Ribonucleoproteína Heterogénea-Nuclear Grupo A-B , Proteínas de Insectos/genética , Proteínas de Unión al ARN/metabolismo , Ribonucleoproteínas/genética , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/metabolismo , Núcleo Celular/química , Mapeo Cromosómico , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Drosophila melanogaster/embriología , Drosophila melanogaster/crecimiento & desarrollo , Técnica del Anticuerpo Fluorescente Indirecta , Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Ribonucleoproteína Nuclear Heterogénea A1 , Ribonucleoproteína Heterogénea-Nuclear Grupo K , Ribonucleoproteínas Nucleares Heterogéneas , Humanos , Proteínas de Insectos/inmunología , Proteínas de Insectos/metabolismo , Datos de Secuencia Molecular , Proteínas Nucleares/metabolismo , Poli C/metabolismo , Pruebas de Precipitina , Unión Proteica , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Transcripción Genética , Vertebrados

RESUMEN

Olfactory adaptation is shown to occur in Drosophila, at both behavioral and physiological levels. In a behavioral paradigm, the extent of adaptation is shown to depend on the dose and duration of the adapting stimulus. Half-maximal adaptation occurred after 15 sec of exposure to an odor, and recovery occurred with a half-time of 1. 5 min, under a set of test conditions. Cross-adaptation was observed among all odor combinations tested, although to a lesser extent than when the same odor was used as both the adapting and the test stimulus. Mutants of the transient receptor potential (Trp) Ca2+ channel were normal in olfactory response, but defective in olfactory adaptation, when measured either behaviorally or in tests of antennal physiology. These results indicate that olfactory response and adaptation can be distinguished. Trp expression was detected in the developing antenna but, surprisingly, not in the mature antenna. These results, together with temperature-shift analysis of a temperature-sensitive trp mutant, provide evidence of a role of Trp in olfactory system development.

Asunto(s)

Adaptación Fisiológica/fisiología , Canales de Calcio/genética , Proteínas de Drosophila , Drosophila/fisiología , Proteínas de Insectos/genética , Neuronas Receptoras Olfatorias/fisiología , Olfato/fisiología , Alelos , Estructuras Animales/química , Animales , Conducta Animal/fisiología , Benzaldehídos , Canales de Calcio/análisis , Canales de Calcio/metabolismo , Electrofisiología , Proteínas de Insectos/análisis , Proteínas de Insectos/metabolismo , Cinética , Mutagénesis/fisiología , Odorantes , Neuronas Receptoras Olfatorias/química , Temperatura , Canales de Potencial de Receptor Transitorio

RESUMEN

The amino acid sequence BCNG-1 (brain cyclic nucleotide gated 1, of the mouse), the first member of mamalian I(h) channels, was used to construct a set of polymerase chain reaction (PCR) primers from possibly conserved regions. Reverse transcription-PCR with Drosophila melanogaster mRNA yielded in a PCR product, which exhibited a high homology to BCNG-1. Using these PCR products to screen a D. melanogaster head cDNA library we isolated a cDNA encoding a member of a new class of putative voltage- and cyclic nucleotide-gated potassium channels from D. melanogaster. The most important features of the amino acid sequence predicted from the cDNA were a C-terminal cyclic nucleotide-binding region, an S4-voltage sensor and a putative potassium-selective pore-forming motif. The high homology of 51% to the sea urchin I(h) channel, which belongs to the same class of ion channels as BCNG-1, leads us to suggest that the Drosophila cDNA is the first insect member of a new class of hyperpolarization-activated and cyclic nucleotide-gated channels. As shown by in situ hybridization, a pronounced mRNA expression was detected in neuronal tissue, including sensory tissue like the compound eyes, and the olfactory and the auditory organs.

Asunto(s)

Canales de Calcio/genética , ADN Complementario , Proteínas de Drosophila , Drosophila melanogaster/genética , Neuronas Aferentes/metabolismo , Canales de Potasio/genética , Homología de Secuencia de Ácido Nucleico , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Canales Catiónicos Regulados por Nucleótidos Cíclicos , Drosophila melanogaster/citología , Ojo/citología , Ojo/metabolismo , Expresión Génica , Genes de Insecto , Hibridación in Situ , Canales Iónicos , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso , Neuronas Aferentes/citología , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

The previously detected ebony (e) locus (Caizzi et al., 1987) consists of a complex gene structure that is divided into seven exons. An open reading frame encoding the putative Ebony protein of 98.5 kDa exhibits homology to a family of peptide synthetases (Stachelhaus and Marahiel, 1995), in good correlation with the proposed function as beta-alanyl-dopamine synthetase. Multiple ebony transcripts are detected throughout development. P-factor mediated transformation of genomic DNA rescues the cuticle, electrophysiological and behavioural phenotypes. Fusion of the ebony reading frame with that of beta-galactosidase of E. coli reveals expression in cuticle and nervous system. Strong staining in the first and, to a lesser extent, in the second optic neuropile may reflect the pronounced visual defect observed in ebony mutants. In addition, weak central brain and thoracic ganglion expression is detected in flies. Conservation of a multidomain protein structure known from peptide synthetases should have functional implications on the putative reaction mechanism of peptide bond formation.

Asunto(s)

Proteínas de Unión al ADN/genética , Proteínas de Drosophila , Drosophila/genética , Genes de Insecto/genética , Sistema Nervioso/metabolismo , Péptido Sintasas/genética , Secuencia de Aminoácidos , Animales , Proteínas Bacterianas/genética , Secuencia de Bases , ADN/química , ADN/genética , ADN/aislamiento & purificación , ADN Complementario , Drosophila/química , Drosophila/crecimiento & desarrollo , Electrorretinografía , Regulación del Desarrollo de la Expresión Génica , Proteínas de Insectos/genética , Larva/química , Larva/enzimología , Datos de Secuencia Molecular , Sistema Nervioso/crecimiento & desarrollo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Análisis de Secuencia de ADN , Transformación Genética , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismo

RESUMEN

Insects perceive a large number of airborne chemicals as olfactory components mainly through the antenna. It is thought that detection of the odorants by specific receptors is followed by a degradative pathway that clears the olfactory organ from accumulating chemicals. In Drosophila, a number of P450 monooxygenases are involved in the metabolism of foreign chemicals [Dunkov et al. (1996) Cytochrome P450 gene cluster in Drosophila melanogaster, Mol. Gen. Genet. 251, 290-297]. NADPH-cytochrome P450 oxidoreductases serve to transfer reducing equivalents to P450 monooxygenases. We isolated cDNA and genomic clones coding for a Drosophila NADPH cytochrome P450 oxidoreductase (CPR). The largest cDNA of 2471 nucleotides in length contained an open reading frame of 693 amino acids that includes the putative CPR sequence. CPR is a single copy gene as shown by genomic Southern hybridisation and maps to the cytogenetic map position 26C on the second chromosome. Comparison of genomic and cDNA CPR sequences revealed a gene structure that is split into at least six exons. The CPR protein sequence is almost identical with that of house fly and remarkably conserved when compared to vertebrates and yeast. RNA expression is high in embryos and antennae as compared to adult heads, adult bodies and larvae. High expression in antennae may reflect the putative function in olfactory clearance.

Asunto(s)

Drosophila melanogaster/enzimología , Drosophila melanogaster/genética , NADPH-Ferrihemoproteína Reductasa/genética , Órganos de los Sentidos/metabolismo , Olfato/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Secuencia Conservada , ADN Complementario/aislamiento & purificación , Drosophila melanogaster/química , Drosophila melanogaster/fisiología , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Genes de Insecto , Proteínas de Insectos/química , Proteínas de Insectos/genética , Proteínas de Insectos/aislamiento & purificación , Proteínas de Insectos/fisiología , Datos de Secuencia Molecular , NADPH-Ferrihemoproteína Reductasa/química , NADPH-Ferrihemoproteína Reductasa/aislamiento & purificación , NADPH-Ferrihemoproteína Reductasa/fisiología , Especificidad de Órganos , Órganos de los Sentidos/fisiología , Olfato/fisiología

RESUMEN

We have isolated cDNAs coding for a ribonucleoprotein of Drosophila melanogaster that is distinguished by its nearly exclusive presence at only one of the several heat shock puffs in polytene chromosomes of third instar larvae. We determined the nucleotide sequence and deduced the corresponding amino acid sequence. Its coding capacity for a 39 kDa protein is consistent with the size of the protein detected by the monoclonal antibody P11 used for expression cloning. Our results show that the P11 protein belongs to the category of hnRNP proteins of bipartite structure: the amino-terminal half contains two RNA binding domains and the carboxyterminal half is rich in glycine residues. Analysis of the genomic structure revealed two introns located within the coding portion of the gene and a third one in the 3'untranslated region. We detect two different polyadenylation sites as a result of alternative termination-polyadenylation. Its strong sequence homology with hnRNP A1 protein and its previously shown association with snRNP particles indicates that a typical hnRNP protein may also exist in a complex with snRNP particles. The P11 sequence corresponds to the Hrb87F sequence that was recently described by Haynes et al. (1) as hnRNP A related gene.

Asunto(s)

Proteínas de Drosophila , Drosophila melanogaster/genética , Ribonucleoproteína Heterogénea-Nuclear Grupo A-B , Proteínas Nucleares , Ribonucleoproteínas/química , Ribonucleoproteínas/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión , Western Blotting , Cromosomas/metabolismo , ADN/genética , Técnica del Anticuerpo Fluorescente , Ribonucleoproteína Nuclear Heterogénea A1 , Ribonucleoproteínas Nucleares Heterogéneas , Intrones , Datos de Secuencia Molecular , ARN/metabolismo , Mapeo Restrictivo , Ribonucleoproteínas/genética , Ribonucleoproteínas Nucleares Pequeñas , Homología de Secuencia de Ácido Nucleico

RESUMEN

Using low stringency hybridisation with a Drosophila melanogaster EF-1 alpha gene fragment we have isolated a genomic DNA clone encoding elongation factor 1 alpha (EF-1 alpha) from Apis mellifera. The hybridising Apis mellifera sequence could be delineated to two small EcoRI fragments that were also revealed by genomic Southern hybridisation. By comparison with the corresponding Drosophila melanogaster data the complete translational reading frame has been deduced. It is interrupted by two intervening sequences of 220 and about 790 nucleotides. Comparison with known eucaryotic EF-1 alpha sequences further confirms that certain amino acid sequences seem to be invariable within the EF-1 alpha protein family.

Asunto(s)

Abejas/genética , ADN/análisis , Drosophila melanogaster/genética , Genes , Factores de Elongación de Péptidos/genética , Secuencia de Aminoácidos , Animales , Abejas/metabolismo , Evolución Biológica , Clonación Molecular , Drosophila melanogaster/metabolismo , Humanos , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Factor 1 de Elongación Peptídica , Biosíntesis de Proteínas , Relación Estructura-Actividad

RESUMEN

A polyclonal antiserum against purified bacteriophage T4 gene 32 protein was raised in rabbits. In Western blots it detected a number of SDS-PAGE separated nuclear and ribosomal proteins of HeLa cells. Using a renaturing blotting system, however, larger hnRNP proteins ranging between 66.000 and 82.000 Da preferentially reacted with this antiserum. In addition hnRNP group A core proteins were detected to a minor extent. Nucleic acid binding proteins like histones or ribosomal proteins were not stained by this antibody after renaturation.

Asunto(s)

Proteínas de Unión al ADN/inmunología , Epítopos/análisis , Ribonucleoproteínas/inmunología , Fagos T/genética , Proteínas Virales/inmunología , Western Blotting , Proteínas de Unión al ADN/aislamiento & purificación , Escherichia coli/genética , Células HeLa/análisis , Ribonucleoproteínas Nucleares Heterogéneas , Humanos , Sueros Inmunes , Peso Molecular , Ribonucleoproteínas/aislamiento & purificación , Proteínas Virales/aislamiento & purificación

RESUMEN

Mouse cDNA clones have been isolated with the help of Drosophila melanogaster 82-kDa heat-shock protein (Hsp82)-coding sequences as hybridization probe. Sequencing of the overlapping mouse clones reveals a long open reading frame (ORF) that encodes a polypeptide of 83.3 kDa which shows about 80% similarity to the respective Drosophila Hsp82 amino acid sequence. The N-terminal half of this cDNA cross-hybridizes to a different class of mouse cDNA clones indicating a related gene. Northern blot hybridization experiments reveal a 2.6-kb poly(A)+RNA when probed with the hsp84 clone and a 2.85-kb signal with the hsp84-related cDNA. The amino acid sequences deduced from the contiguous ORF of the hsp84 and the hsp84-related cDNA coincide with the N-terminal sequence of formerly identified 84-kDa and 86-kDa tumour-specific transplantation antigens (Ullrich et al., 1986). In addition, the amino acid composition of the putative 84-kDa mouse Hsp described here is very similar to that of the 84-kDa tumour antigen described by Ullrich et al. (1986). Both observations corroborate the assumption that these Hsps are identical to the described 84-kDa and 86-kDa tumour-specific transplantation antigens. Using these mouse hsp gene clones as hybridization probes we also isolated the corresponding pair of human cDNA clones. Comparison of the respective sequences reveals a strong evolutionary constraint on these two genes in mouse and man.

Asunto(s)

Antígenos de Neoplasias/genética , Proteínas de Choque Térmico/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Northern Blotting , Southern Blotting , Clonación Molecular , Drosophila melanogaster , Pruebas Genéticas , Humanos , Ratones , Datos de Secuencia Molecular , Peso Molecular , Familia de Multigenes , ARN/análisis

RESUMEN

Tyrosine sulfation is a post-translational modification in the trans Golgi that has been found in all animal species studied. In the preceding paper (Baeuerle, P. A., Lottspeich, F., and Huttner, W. B. (1988) J. Biol. Chem. 263, 14925-14929), we have identified the site of tyrosine sulfation in an insect secretory protein, yolk protein 2 (YP2) of Drosophila melanogaster. In the present report, tyrosine sulfation of this protein was examined after expression in a heterologous mammalian cell system. Mouse fibroblasts, transfected with Drosophila YP2 genomic DNA inserted into the eucaryotic expression vector pSV2, secreted the fly protein in sulfated form. Analyses of Drosophila YP2 produced by the mouse cells showed that the features of sulfation of this protein were identical to those previously determined for YP2 isolated from flies. YP2 secreted from mouse fibroblasts was found to be exclusively sulfated on tyrosine residues. The stoichiometry of tyrosine sulfation was approximately 1 mol of sulfate/mol of YP2. Sulfate was linked to the same tyrosine residue as in YP2 isolated from flies, tyrosine 172. These results show that essential parameters of the tyrosine sulfation reaction are very similar in insects and mammals and thus highly conserved in evolution.

Asunto(s)

Drosophila melanogaster/genética , Proteínas del Huevo/genética , Genes , Procesamiento Proteico-Postraduccional , Transfección , Animales , Línea Celular , ADN Recombinante/metabolismo , Proteínas del Huevo/biosíntesis , Técnica del Anticuerpo Fluorescente , Immunoblotting , Células L/metabolismo , Ratones , Plásmidos , Sulfatos/metabolismo , Radioisótopos de Azufre , Ácidos Sulfúricos/metabolismo , Tirosina

RESUMEN

We have characterized two previously cloned genes, F1 and F2 (1) that code for elongation factor EF - 1 alpha of Drosophila melanogaster. Genomic Southern blot hybridization revealed that they are the only gene copies present. We isolated cDNA clones of both transcripts from embryonal and pupal stage of development that cover the entire transcription unit. The 5' ends of both genes have been determined by primer extension and for F1 also by RNA sequencing. These start sites have been shown to be used consistently during development. Comparison of cDNA and genomic sequences revealed that EF - 1 alpha,F1 consists of two and EF - 1 alpha,F2 of five exons. The two described elongation factor genes exhibit several regions of strong sequence conservation when compared to five recently cloned eucaryotic elongation factors.

Asunto(s)

Drosophila melanogaster/genética , Factores de Elongación de Péptidos/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , ADN/genética , Drosophila melanogaster/crecimiento & desarrollo , Regulación de la Expresión Génica , Genes , Datos de Secuencia Molecular , Factor 1 de Elongación Peptídica , Regiones Promotoras Genéticas , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie

Asunto(s)

Regulación de la Expresión Génica , Proteínas de Choque Térmico/biosíntesis , Calor , Factores de Elongación de Péptidos/biosíntesis , Animales , ADN/genética , Proteínas de Choque Térmico/genética , Células L/análisis , Ratones , Factor 1 de Elongación Peptídica , Factores de Elongación de Péptidos/genética , Ratas , Proteínas Recombinantes/biosíntesis

RESUMEN

A comparison of gene structure, sequence, and transcription pattern of heat shock loci 93D of Drosophila melanogaster and 48B of Drosophila hydei has been performed. Both heat shock loci consist of an unique region that is flanked by an internally repetitive element. Different members of these elements are highly conserved, repeat unit length, however, and primary sequence diverged totally. Whereas the overall gene structure in both species is substantially related, sequence conservation is only observed at very few sites in the unique region. These represent primarily sequences that are identified as regulatory elements for faithful transcription and processing. The number and size of transcripts obtained from heat shock locus 48B in third instar larvae closely resembles the pattern of heat shock locus 93D. Thus their quite alike structure and transcription pattern suggest strongly a conserved hitherto unknown function.

Asunto(s)

Drosophila melanogaster/genética , Drosophila/genética , Proteínas de Choque Térmico/genética , Animales , Genes , Hibridación de Ácido Nucleico , Secuencias Repetitivas de Ácidos Nucleicos , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Transcripción Genética

RESUMEN

Acetylcholine is a major excitatory neurotransmitter in the central nervous system of insects. Using DNA probes of the Torpedo nicotinic acetylcholine receptor (AChR) we have isolated two overlapping cDNA clones encoding a putative neuronal AChR protein from the fruitfly, Drosophila melanogaster. The predicted mature protein consists of 497 amino acids, has a calculated mol. wt of 57 340 and shows extensive homology to known AChR subunits from different species along its entire amino acid sequence. Northern analysis revealed a hybridizing mRNA of 3.2 kb in late embryo and in pupae. Expression of the corresponding AChR gene thus characterizes periods of neuronal differentiation in Drosophila.

RESUMEN

Screening a cDNA library of 2- to 3-day-old flies with poly(A)(+) RNA from male and female flies, we were able to isolate a small number of clones hybridizing preferentially with RNA from female flies. Four cDNA clones, derived from a single mRNA species, proved to be highly abundant in female flies. When we screened a genomic library with the longest cDNA, we obtained two genomic clones, F1 and F2; F1 was a direct copy of the cDNA and F2 was obtained by cross-hybridization. A detailed analysis of these genomic clones revealed two independent genes coding for proteins of 50 kDa that are >90% homologous. RNA analysis with gene-specific probes from the 3' untranslated region showed an expression of F1 in all stages of development with a 5- to 10-fold overexpression of this RNA in female flies compared with males. In contrast to F1, F2 is mainly expressed in late pupae and is expressed only at low levels in adult flies.

RESUMEN

The heat-shock locus 93D from Drosophila melanogaster has recently become available for a molecular analysis. We established a restriction site map of a recombinant DNA clone covering the major part of heat-shock locus 93D. This clone includes part of a repetitive Taq I region and neighbouring unique sequences. The portion of the Taq I repeat analysed consists of tandemly arranged sequence blocks of about 280 base pairs (bp) in length. Using genomic and cDNA as hybridization probes we examined the transcription of 93D in 2- to 4-day-old flies. We identified two major RNA classes enhanced after heat shock, namely nonpolyadenylated transcripts of heterogeneous length derived from the repetitive region and one discrete polyadenylated transcript in spliced and unspliced form from the neighbouring unique region. The occurrence of a highly heterogeneous poly(A)- transcript and high levels of an unspliced discrete poly(A)+ species suggests unusual mechanisms of transcription regulation in the 93D region.

Asunto(s)

Drosophila melanogaster/genética , Proteínas de Choque Térmico/genética , ARN/genética , Transcripción Genética , Animales , Clonación Molecular , ADN/metabolismo , Enzimas de Restricción del ADN , ADN Recombinante/metabolismo , Genes , Hibridación de Ácido Nucleico

RESUMEN

We have isolated and characterized a recombinant which contains a Drosophila melanogaster U1 small nuclear RNA (snRNA) gene colinear with the published snRNA sequence. Southern hybridizations of the fly genomic DNA, using as probe a plasmid containing only the coding region of the gene, shows that the fly contains at most three or four genes and very few related sequences for the small nuclear U1 RNA. These genes were localized by in situ hybridization at different chromosomal loci and show no spatial relationship to the U2 snRNA genes.

Asunto(s)

Genes , ARN/genética , Animales , Autorradiografía , Secuencia de Bases , Mapeo Cromosómico , Enzimas de Restricción del ADN , Drosophila melanogaster/genética , Hibridación de Ácido Nucleico , ARN Nuclear Pequeño

RESUMEN

Four different U2-snRNA genes/related sequences of D. melanogaster were cloned and characterized. The sequences of all four genes suggest that they were generated by a DNA-mediated mechanism. These genes/related sequences were found to be located in two loci, each locus containing two U2 snRNA sequences. Using coding sequences as well as flanking sequences as hybridization probes against polytene chromosomes of D. melanogaster Oregon R we were able to map these loci separately at positions 34BC and 84C. By Northern analysis we observed that the quantities of U2- and U1-snRNA are coordinated and change during the embryonic development of the fly.

Asunto(s)

Drosophila melanogaster/genética , ARN/genética , Animales , Secuencia de Bases , Mapeo Cromosómico , Drosophila melanogaster/crecimiento & desarrollo , Regulación de la Expresión Génica , Genes , Hibridación de Ácido Nucleico , ARN/biosíntesis , ARN Nuclear Pequeño

RESUMEN

Using the microcloning approach a number of recombinant lambda phages carrying DNA from the 93D region have been isolated. Screening genomic libraries, cloned in phage lambda or cosmid vectors, with this isolated DNA yielded a series of overlapping DNA fragments from the region 93D6-7 as shown by in situ hybridization to polytene chromosomes. In vitro 32P-labelled nuclear RNA prepared from heat-shocked third instar larvae hybridized specifically to one fragment within 85 kb of cloned DNA. The region which is specifically transcribed after heat shock could be defined to a cluster of internally-repetitive DNA and its neighbouring proximal sequences. Over a sequence of 10-12 kb in length the DNA is cut into repeat units of approximately 280 nucleotides by the restriction endonuclease TaqI. The TaqI repeat sequences are unique in the Drosophila genome.

Asunto(s)

Clonación Molecular , Drosophila melanogaster/genética , Genes , Proteínas de Choque Térmico/genética , Animales , Secuencia de Bases , Enzimas de Restricción del ADN , Peso Molecular , Hibridación de Ácido Nucleico , Plásmidos , Secuencias Repetitivas de Ácidos Nucleicos , Transcripción Genética