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PubMed1996

Increased specificity of colloidal silver staining by means of chemical attenuation.

A modification of the silver staining procedure of Howell and Black (1980) is reported which makes use of teleostean gelatin as protective colloid and renders a high signal-to-noise ratio. We demonstrate that this ratio can be further increased by subsequent attenuation with a chemical reducer consisting of a mixture of potassium ferricyanide (III) and sodium thiosulphate. It is shown that slight changes of the concentration of the reactive compounds of the chemical reducer make the protocol applicable to human, plant (Aliium cepa, Rhinanthus minor) as well as meiotic insect (Acheta domesticus) chromosome preparations. Due to its broad applicability, the method could find utilization in studies on chromatin and chromosome functions in many species.

Animals↗
PubMed2001

Genome analysis: More Drosophila Y chromosome genes.

The Drosophila melanogaster Y chromosome has long been known to contain few functional genes other than several required for male fertility. The D. melanogaster genome sequence has now allowed characterization of two more male fertility genes, shedding light on the function and evolution of Y chromosomes.

Animals↗
PubMed1978

Messenger RNA for the insect storage protein calliphorin: in vitro translation and chromosomal hybridization analyses of a 20 S poly(A)-RNA fraction.

A major poly(A)-containing RNA fraction of the approximate size expected of a monocistronic mRNA for the storage protein calliphorin has been isolated from the larval fat bodies of Calliphora vicina during early instar 3. This 20 S RNA fraction programs the synthesis by cell-free wheat embryo extracts of polypeptides of 86,000 daltons identified by tryptic peptide fingerprinting as precursors of the authentic calliphorin subunits of 83,000 daltons. Complementary DNA synthesized by AMV reverse transcriptase using the same 20 S RNA as template hybridized in situ to a single segment of one or two bands in the salivary polytene chromosomes of C. vicina.

Adipose Tissue↗
PubMed2003

Concurrence of losing a chromosome and the ability to produce destruxins in a mutant of Metarhizium anisopliae.

In a previous study, a spontaneous subtilisin pr1A and pr1B gene-deficient mutant of the entomopathogenic fungus Metarhizium anisopliae strain V275 has been identified [Wang, C.-S. et al. (2002) FEMS Microbiol. Lett. 213, 251-255]. The insecticidal metabolites of this mutant were studied further. High-performance liquid chromatography (HPLC) analysis indicated that the mutant isolate lost the ability to produce cyclic peptide toxins, destruxins, both in vitro and in vivo. Pulsed-field gel electrophoresis revealed that the mutant concurrently lost a 1.05 Mb (approximately) chromosome, demonstrating for the first time that a conditionally dispensable (CD) chromosome exists in the insect pathogenic fungus, M. anisopliae. Concurrence of losing the ability to produce destruxins and a CD chromosome in the mutant suggests that the toxin synthetase genes of M. anisopliae are located on this CD chromosome, as similarly described for plant pathogenic fungi. Semi-quantitative api ZYM analysis showed more biochemical disparities between the mutant and the wild-type strain.

Acetylglucosaminidase↗
PubMed1996

Cytotaxonomical examination for sibling species in the taxon Anopheles culicifacies Giles in Sri Lanka.

Ovarian polytene chromosomes of Anopheles culicifacies collected from different climatic and eco-epidemiological localities were examined to study the composition and distribution of sibling species in Sri Lanka. During the study period 4328 ovaries were prepared for examination, of which 1937 were suitable for the reading of polytene chromosomes. Among these 458 specimens were identified as belonging to species B based on diagnostic inversion on x-chromosome and 1479 definitively were identified as sibling species B on the basis of x-chromosome and chromosome arm-2 inversion. Other sibling species were not encountered among the specimens identified from 31 Health Areas during the study period. The occurrence of sibling species B of An. culicifacies in Sri Lanka is confirmed but no evidence of other sibling species (A, C or D) was found.

Animals↗
PubMed1994

Structure and chromosomal localization of CYP6A1, a cytochrome P450-encoding gene from the house fly.

We determined the sequence of a cytochrome P450-encoding gene, CYP6A1, in an insecticide-resistant strain (Rutgers) and in insecticide-susceptible strains (aabys and sbo) of the house fly Musca domestica. The deduced amino acid (aa) sequence of CYP6A1 is 98% identical between Rutgers and aabys, and it is identical between Rutgers and sbo. Differences in aa sequence occur in regions that are not thought to participate in the active site. CYP6A1 has a single intron, 60 bp in length. It does not occur in a position homologous to intron sites in genes of the related CYP3 family or other vertebrate P450 families. We mapped the transcription start point (tsp), TATA box and barbie box [Shaw and Fulco, J. Biol. Chem. 269 (1993) 2997-3004] of this phenobarbital-inducible gene. Although overexpression of CYP6A1 in strain Rutgers is controlled by a locus on chromosome II [Cariño et al., Insect Biochem. Mol. Biol. 24 (1994) 411-418], we have localized CYP6A1 to chromosome V, thus showing that a trans-acting factor controls the overexpression of the gene in the Rutgers strain.

Amino Acid Sequence↗
PubMed2001

Genetic and physical mapping in mosquitoes: molecular approaches.

The genetic background of individual mosquito species and populations within those species influences the transmission of mosquito-borne pathogens to humans. Technical advances in contemporary genomics are contributing significantly to the detailed genetic analysis of this mosquito-pathogen interaction as well as all other aspects of mosquito biology, ecology, and evolution. A variety of DNA-based marker types are being used to develop genetic maps for a number of mosquito species. Complex phenotypic traits such as vector competence are being dissected into their discrete genetic components, with the intention of eventually using this information to develop new methods to prevent disease transmission. Both genetic- and physical-mapping techniques are being used to define and compare genome architecture among and within mosquito species. The integration of genetic- and physical-map information is providing a sound framework for map-based positional cloning of target genes of interest. This review focuses on advances in genome-based analysis and their specific applications to mosquitoes.

Animals↗
PubMed1999

Genomic imprinting and position-effect variegation in Drosophila melanogaster.

Genomic imprinting is a phenomenon in which the expression of a gene or chromosomal region depends on the sex of the individual transmitting it. The term imprinting was first coined to describe parent-specific chromosome behavior in the dipteran insect Sciara and has since been described in many organisms, including other insects, plants, fish, and mammals. In this article we describe a mini-X chromosome in Drosophila melanogaster that shows genomic imprinting of at least three closely linked genes. The imprinting of these genes is observed as mosaic silencing when the genes are transmitted by the male parent, in contrast to essentially wild-type expression when the same genes are maternally transmitted. We show that the imprint is due to the sex of the parent rather than to a conventional maternal effect, differential mitotic instability of the mini-X chromosome, or an allele-specific effect. Finally, we have examined the effects of classical modifiers of position-effect variegation on the maintenance and the establishment of the imprint. Factors that modify position-effect variegation alter the somatic expression but not the establishment of the imprint. This suggests that chromatin structure is important in maintenance of the imprint, but a separate mechanism may be responsible for its initiation.

Alleles↗
PubMed1965

THE SALIVARY GLAND CHROMOSOMES OF ANOPHELES OCCIDENTALIS.

Increasing emphasis is being placed on the study of the genetics of insect vectors of disease, and, although Anopheles occidentalis is not itself a malaria vector, study of its salivary gland chromosomes is important as part of investigations into the phylogenetic and evolutionary relationships of anophelines, especially in view of this species' close morphological similarity to other members of the A. maculipennis complex.The authors of this paper provide a map of the salivary chromosomes of A. occidentalis, and discuss the many points of resemblance of those chromosomes to those of A. freeborni in particular. However, they point out that at least ten different paracentric inversions must be postulated to derive the A. occidentalis chromosomes from those of A. freeborni.

Animals↗
PubMed2000

The effect of 1,10-phenanthroline on the chromosome damage and sister-chromatid exchanges induced by streptozotocin in mammalian and insect cells.

The effect of the metal chelating agent 1,10-Phenanthroline (PNT) on the streptozotocin (STZ)-induced chromosomal aberrations (CAs) and sister-chromatid exchanges (SCEs) in Chinese hamster ovary (CHO) and mosquito (Aedes albopictus) cells was investigated. Treatment of CHO and mosquito cells with STZ produced a significant and dose-response increase in the yield of CAs as well as SCEs (p<0.05). The addition of PNT prevented the induction of CAs by STZ in both types of cells, causing a significant decrease in the frequency of STZ-induced CAs (46.5-72.5%) (p<0.05). This fact indicates that intracellular transition metals are implicated in STZ-induced CAs and that the Fenton reaction (Fe(2+)+H(2)O(2)-->OH degrees +OH(-)+ Fe(3+)) is partly responsible for the production of CAs by this compound. On the other hand, the addition of PNT to CHO and mosquito cell cultures did not prevent the induction of SCEs by STZ. Therefore, it is valid to assume that the induction of CAs and SCEs by STZ occurs by different mechanisms.

Animals↗
PubMed2003

The origin of the achiasmatic XY sex chromosome system in Cacopsylla peregrina (Frst.) (Psylloidea, Homoptera).

The status of an extra univalent, if it is a B chromosome or an achiasmatic Y chromosome, associating with the X chromosome in male meiosis of Cacopsylla peregrina (Frst.) (Homoptera, Psylloidea) was analysed. One extra univalent was present in all males collected from three geographically well separated populations, it was mitotically stable, and showed precise segregation from the X chromosome. These findings led us to propose that the univalent represents in fact a Y chromosome. The behaviour of the X and Y chromosomes during meiotic prophase suggested that their regular segregation was based on an achiasmatic segregation mechanism characterised by a 'touch and go' pairing of segregating chromosomes at metaphase I. To explain the formation of the achiasmatic Y within an insect group with X0 sex chromosome system, it was suggested that the Y chromosome has evolved from a mitotically stable B chromosome that was first integrated into an achiasmatic segregation system with the X chromosome, and has later become fixed in the karyotype as a Y chromosome.

Animals↗
PubMed1983

Complete dependence between AG NORs and C-positive heterochromatin revealed by simultaneous AG-NOR C-banding method.

In the present paper we report a technique which permits the simultaneous observation of C-banded material and the active NORs or nucleoli in both mitotic and meiotic chromosomes of mammals and insects. The use of the present technique permits to establish a complete correlation between the C-banded positive regions and the presence of nucleoli associated to them. Some aspects related with this correspondence are discussed.

Animals↗
PubMed2006

Molecular and biological characterization of deformed wing virus of honeybees (Apis mellifera L.).

Deformed wing virus (DWV) of honeybees (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. The virus was purified from diseased insects, and its genome was cloned and sequenced. The genomic RNA of DWV is 10,140 nucleotides in length and contains a single large open reading frame encoding a 328-kDa polyprotein. The coding sequence is flanked by a 1,144-nucleotide 5' nontranslated leader sequence and a 317-nucleotide 3' nontranslated region, followed by a poly(A) tail. The three major structural proteins, VP1 (44 kDa), VP2 (32 kDa), and VP3 (28 kDa), were identified, and their genes were mapped to the N-terminal section of the polyprotein. The C-terminal part of the polyprotein contains sequence motifs typical of well-characterized picornavirus nonstructural proteins: an RNA helicase, a chymotrypsin-like 3C protease, and an RNA-dependent RNA polymerase. The genome organization, capsid morphology, and sequence comparison data indicate that DWV is a member of the recently established genus Iflavirus.

Amino Acid Sequence↗
PubMed1994

Assembly and disassembly of spliceosomes along a specific pre-messenger RNP fiber.

Transcriptionally active Balbiani ring (BR) genes in the salivary glands of the dipteran Chironomus tentans were studied by immunoelectron microscopy to establish the distribution of spliceosome components along a specific pre-messenger ribonucleoprotein (pre-mRNP) fiber. The BR genes are 35-40 kb in size with three introns close to the 5' end and one close to the 3' end; a very large middle portion lacks introns. As a rule the 5' introns are spliced concomitant with transcription in the promoter proximal third of the gene, while the 3' intron is spliced post-transcriptionally. The BR genes with growing pre-mRNPs were visualized in situ, while completed and released pre-mRNPs were isolated from the nucleoplasm and studied unfolded on a grid surface. An anti-snRNP antibody (Y12) bound mainly to the promoter proximal third of the BR gene (86%) and only to a minor extent to the middle and distal thirds (7 and 7% respectively). An antibody to an hnRNP protein reacted with the proximal, middle and distal regions to an increasing extent (17, 38 and 45% respectively), reflecting the increase in size of the growing transcription product. In the nucleoplasmic pre-mRNP particle only one end of the RNP fiber was labeled by Y 12, presumably the 3' end; the anti-hnRNP antibody decorated the entire RNP fiber. Thus, the snRNPs do not associate along the whole pre-mRNP fiber but rather bind to the 5' and 3' ends, i.e. the regions containing the introns. The results also imply that the spliceosomes both assemble and disassemble rapidly on the pre-mRNP fiber.

Animals↗
PubMed2006

Fine-scale crossover rate heterogeneity in Drosophila pseudoobscura.

Broad-scale differences in crossover rate across the genome have been characterized in most genomes studied. Fine-scale differences, however, have only been examined in a few taxa, such as Arabidopsis, yeast, humans, and mice. No prior studies have directly looked for fine-scale recombination rate heterogeneity in Drosophila. We produced 370 Drosophila pseudoobscura containing a crossover event within the 2-megabase (MB) region between the genes yellow and white. We then examined 19 intervals within this region and determined where the crossovers occurred. We found that recombination events occur nonrandomly on a small scale and that mild "hotspots" of a few kilobases exist in Drosophila. Among the regions studied, recombination rates varied from 1.4 to 52 cM/MB. We also observed a trend toward high codon bias in regions of high recombination. Finally, we identified a significantly positive correlation between recombination rate and simple repeats, as well as the motif CACAC. These sequence features may contribute to broad-scale variation in crossover rate and, thus, shed light on features associated with crossover rate heterogeneity at a genome-wide scale.

Animals↗