C banding in polytene chromosomes of Simulium ornatipes and S. melatum (Diptera: Simuliidae)

Polytene and mitotic chromosomes of Simulium ornatipes and S. melatum were subjected to C banding procedures. In both species polytene chromosomes consistently show C banding of centromere regions, telomeres, nucleolar organiser and, unexpectedly, numerous interstitial sites. The interstitial C banding sites correspond to morphologically single polytene bands. Their response is graded and independent of band size. Interstitial C bands in S. ornatipes are scattered throughout the complement, whereas in S. melatum they are clustered. Supernumerary heterochromatic segments in S. ornatipes also exhibit strong C banding and inverted segments can differ from standard in C banding pattern. — Mitotic chromosomes of both species show a single centric C band with indications of two weak interstitial bands in S. ornatipes, suggesting that many C band regions, detectable in polytene chromosomes, are not resolved by present techniques in mitotic chromosomes. — Contrary to current opinion that C banding is diagnostic for constitutive heterochromatin, the interstitial C band sites of polytene chromosomes are regarded as euchromatic. Conversely, the heterochromatic pericentric regions of S. ornatipes are not C banded. — It appears that polytene chromosomes offer a promising system for the elucidation of C banding mechanisms.     

Polytene chromosomes in pupal and adult blackflies (Diptera: Simuliidae)

A number of pupal and adult tissues of eight Australian blackfly species representing three genera, Austrosimulium, Cnephia and Simulium, were examined for the presence of polytene chromosomes. Banded polytene chromosomes were found in malpighian tubules, hind gut, fat body, and ovary, but only those from the malpighian tubules of female adults and pupae were of good quality. A detailed comparison of polytene chromosomes from larval salivary glands and adult malpighian tubules was made in S. ornatipes and, to a limited extent, in S. melatum. The banding patterns of chromosomes from both tissues were found to be identical with minor differences in puffing patterns in S. ornatipes and chromocenter characteristics in S. melatum. A survey of the remaining six species shows five of them to have malpighian chromosomes suitable for detailed cytological analysis. Simultaneous studies of larval, pupal and adult polytene chromosome systems offer a novel approach to the analysis of population problems in blackflies. The ability to recognise sibling species in adults also has potential practical significance in efforts to control vectors of onchocerciasis.     

Karyotype structure in species of Propsilocerus genus (Diptera, Chironomidae)

Karyotype structure and polytene chromosome banding patterns were studied in two Orthocladiinae siblings--Propsilocerus akamusi (China) and Propsilocerus jacuticus (Russia). Both species have haploid number of chromosome typical for Orthocladiinae (n = 3). An unusual structure of centromeric regions was observed in all three chromosomes of karyotypes in both species. Photomaps of polytene chromosomes are presented. A comparison of karyotypes of P. akamusi and Propsilocerus jacuticus revealed a high level of homology in their banding sequences, however, the presence of fixed paracentric inversions in chromosomal arms IR, IIR, IIIR of Propsilocerus jacuticus has shown a clear-cut phylogenetic divergence. No chromosomal polymorphism was found in both species.     

Identical functional organization of nonpolytene and polytene chromosomes in Drosophila melanogaster

Salivary gland polytene chromosomes demonstrate banding pattern, genetic meaning of which is an enigma for decades. Till now it is not known how to mark the band/interband borders on physical map of DNA and structures of polytene chromosomes are not characterized in molecular and genetic terms. It is not known either similar banding pattern exists in chromosomes of regular diploid mitotically dividing nonpolytene cells. Using the newly developed approach permitting to identify the interband material and localization data of interband-specific proteins from modENCODE and other genome-wide projects, we identify physical limits of bands and interbands in small cytological region 9F13-10B3 of the X chromosome in D. melanogaster, as well as characterize their general molecular features. Our results suggests that the polytene and interphase cell line chromosomes have practically the same patterns of bands and interbands reflecting, probably, the basic principle of interphase chromosome organization. Two types of bands have been described in chromosomes, early and late-replicating, which differ in many aspects of their protein and genetic content. As appeared, origin recognition complexes are located almost totally in the interbands of chromosomes.     

Characteristics of inserted mutations obtained in the P-M hybrid dysgenesis system in the 9F12-10A7 region of the Drosophila melanogaster X-chromosome

19 new mutations in the 9F12-10A7 region of Drosophila melanogaster X chromosome was obtained in the system of P-M hybrid dysgenesis. They appeared to be lethals, as judged from viability of homo- or hemizygous females. In situ hybridization of P DNA with polytene chromosomes revealed P-element insertion in the 10A1-2 band in the majority of the mutants. As a result of complementation analysis, all these mutations were localized at previously known loci: l(1)BP1, l(1)BP5, l(1)BP8, l(1)BP7. No insertion mutations were found at the vermilion locus. This can imply for non-random distribution of insertion mutations in the region studied. Further comparison of these mutations with previously EMS-induced ones revealed that insertion mutations are predominantly hypomorph lethals which do not influence the viability, morphology and fertility of homozygous males and females, but drastically reduce viability of hemizygous females.     

Chromosomal organization of Drosophila tumours

In otu mutants of Drosophila melanogaster ovarian tumours develop because of the high mitotic activity of the mutant cystocytes; the latter are normally endopolyploid. In certain alleles of otu, however, a varying proportion of the mutant ovarian cystocytes undergo polyteny. Mutant cystocytes with polytene chromosomes are termed pseudonurse cells (PNC). Polytene chromosome morphology and banding patterns in PNC of otu ¹/otu³ flies were cytologically analysed. Extensive variability was noted in the quality of the banding pattern of the PNC chromosomes which ranged from highly condensed (condensed PNC chromosomes) to those with a banding pattern (banded PNC chromosomes) similar to that in larval salivary gland cells (SGC). Both the condensed and banded PNC chromosomes frequently enter into a diffuse state characterised by weakened synapsis of the polytene chromatids and alterations in their banding pattern (diffuse PNC chromosomes). Analysis of DNA synthesis patterns in the various morphological forms of PNC polytene chromosomes by ³H-thymidine autoradiography revealed a basic similarity to the pattern seen in polytene nuclei of larval SGC. Independently replicating sites, however, could be unambiguously identified only in banded PNC chromosomes. Comparison of late replicating sites in such PNC chromosomes with those of larval SGC showed a remarkable similarity in the two cell types. These results suggest a close correlation between the polytene chromosome banding pattern and its replicative organization.     

Cytogenetic variation in natural populations of the Old World screwworm fly Chrysomya bezziana (Diptera: Calliphoridae).

The existence of sibling species in the Old World screwworm fly Chrysomya bezziana would raise serious problems in eradicating this pest if it entered Australia. Cytogenetic variation in C. bezziana was investigated by analyzing pupal trichogen polytene chromosomes. Natural populations of C. bezziana spanning its range from southern Africa to Papua New Guinea were examined as well as hybrids between a New Guinea laboratory strain and natural populations. No evidence of sibling species was found. All populations exhibited the same basic banding pattern as the standard sequence established from a Papua New Guinea strain. Extensive asynapsis of chromosome homologues was found in some hybrid crosses and was therefore measured in all populations and hybrids to detect systematic variation. Asynapsis levels in most hybrids could not be statistically distinguished from those present in the parent populations except for crosses between populations at the ends of the range. This result does not permit asynapsis levels to be used in establishing the origin of introduced flies by estimating their distance from known populations. One inversion polymorphism and six band polymorphisms spread over three chromosomes were analyzed. Populations in each sampled region had characteristic combinations of band polymorphisms. This may offer a diagnostic method for determining the origin of flies accidentally introduced to Australia.     

Sequence replication and banding organization in the polytene chromosomes of Drosophila melanogaster

The relative proportions of cloned DNA fragments from all known hierarchies of sequence organization in polytene and diploid chromosomes were compared. It was found that unique sequences of varying sizes and chromosomal locations are equally replicated in salivary gland chromosomes. Sequences of euchromatic polydisperse gene families are also replicated proportionately in polytene and diploid tissues. Perhaps the most significant finding is that the histone gene repeats, despite their normal banding organization, are under-replicated in the polytene chromosome of Drosophila melanogaster. However, the clustered and well-banded 5S genes are most likely equally replicated. It is therefore concluded that differential sequence replication plays no apparent role in either the assembly or morphology of a band; and likewise, the assembly of polytenic DNA into band units is not affected by either the local abundancy or arrangement of middle repetitive sequences. The likelihood that the clustered arrangement is an important factor in the selection of sequences for under-replication is discussed.     

Polytene chromosomes of the Old World screwworm fly (Chrysomya bezziana) and its evolutionary relationships with Lucilia cuprina and Cochliomyia hominivorax (Diptera: Calliphoridae).

Standard polytene chromosome maps for the Old World screwsworm fly, Chrysomya bezziana, are presented. Good quality polytene chromosomes obtainable from pupal trichogen cells allow detailed analysis of autosomal euchromatin. The sex chromosomes are represented by irregular heterochromatic structures resembling those described previously in trichogen polytene chromosomes of the Australian sheep blowfly, Lucilia cuprina. A high degree of homology with the banding pattern of L. cuprina polytene chromosomes allowed direct recognition of approximately 60% of the L. cuprina complement in the C. bezziana maps. A further 13% may be homologous. The extensive homology observed is discussed in relation to the rate of chromosome rearrangement and conservation of karyotype elements in the evolution of Calliphorid flies. The observed conservation in polytene banding patterns should facilitate construction of phylogenies over a number of generic groups.     

Maintenance of chromosome arm integrity between two Anopheles mosquito subgenera

Low-resolution chromosomal homology between Anopheles gambiae and A. albimanus was determined by polytene chromosome in situ cross hybridization of 17 recombinant DNA and PCR products hybridizing to 23 loci. Hybridization results reflect that the chromosomes have rearranged in the form of autosomal whole-arm translocations and numerous paracentric inversions and not by large detectable pericentric inversions or partial arm translocations. An. gambiae and An. albimanus chromosomes hence differ from each other by possessing alternative autosomal arm associations and rearranged internal structure of each arm, but the integrity of the whole arms has remained conserved. In addition, a photomap of the larval salivary gland polytene chromosomes of An. albimanus that we used to identify sites of hybridization in this species is presented that delineates further banding details than maps published in the past.     

Genetic and cytogenetic analysis of the American cherry fruit fly, Rhagoletis cingulata (Diptera: Tephritidae)

The American eastern cherry fruit fly, Rhagoletis cingulata, a pest of cherries in the western hemisphere, invaded Europe in 1983, and since then dispersed to several European countries. Information on the genetics and cytogenetics of this pest is very scarce. The mitotic karyotype and detailed photographic maps of the salivary gland polytene chromosomes of R. cingulata are presented here. The mitotic metaphase complement consists of six pairs of chromosomes with the sex chromosomes being very small and similar in size. The analysis of the salivary gland polytene complement shows a total number of five long chromosomes (10 polytene arms), which correspond to the five autosomes of the mitotic nuclei and an extrachromosomal heterochromatic mass, which corresponds to the sex chromosomes. The banding patterns and the most characteristic features and prominent landmarks of each polytene chromosome are presented and discussed. Chromosomal homologies between R. cingulata, R. completa and R. cerasi are also proposed, based on the comparison of chromosome banding patterns. Furthermore, the detection and characterization of Wolbachia pipientis in the R. cingulata population studied is presented and the potential correlation with the asynaptic phenomena found in its polytene complement is discussed. In addition, 10 out of 24 microsatellite markers developed for other Rhagoletis species are cross-amplified, evaluated and proposed as useful markers for population and genetic studies in R. cingulata.     

Banding patterns in Drosophila melanogaster polytene chromosomes correlate with DNA-binding protein occupancy

The most enigmatic feature of polytene chromosomes is their banding pattern, the genetic organization of which has been a very attractive puzzle for many years. Recent genome-wide protein mapping efforts have produced a wealth of data for the chromosome proteins of Drosophila cells. Based on their specific protein composition, the chromosomes comprise two types of bands, as well as interbands. These differ in terms of time of replication and specific types of proteins. The interbands are characterized by their association with "active" chromatin proteins, nucleosome remodeling, and origin recognition complexes, and so they have three functions: acting as binding sites for RNA pol II, initiation of replication and nucleosome remodeling of short fragments of DNA. The borders and organization of the same band and interband regions are largely identical, irrespective of the cell type studied. This demonstrates that the banding pattern is a universal principle of the organization of interphase polytene and non-polytene chromosomes.     

Genome re-assignment of Arachis trinitensis (Sect. Arachis, Leguminosae) and its implications for the genetic origin of cultivated peanut

The karyotype structure of Arachis trinitensis was studied by conventional Feulgen staining, CMA/DAPI banding and rDNA loci detection by fluorescence in situ hybridization (FISH) in order to establish its genome status and test the hypothesis that this species is a genome donor of cultivated peanut. Conventional staining revealed that the karyotype lacked the small "A chromosomes" characteristic of the A genome. In agreement with this, chromosomal banding showed that none of the chromosomes had the large centromeric bands expected for A chromosomes. FISH revealed one pair each of 5S and 45S rDNA loci, located in different medium-sized metacentric chromosomes. Collectively, these results suggest that A. trinitensis should be removed from the A genome and be considered as a B or non-A genome species. The pattern of heterochromatic bands and rDNA loci of A. trinitensis differ markedly from any of the complements of A. hypogaea, suggesting that the former species is unlikely to be one of the wild diploid progenitors of the latter.     

Physical mapping of 5S and 18S-26S ribosomal RNA gene families inAllium victorialis var.platyphyllum

A physical map of the 5S and 18S–26S rRNA genes was determined using bi-color fluorescencein situ hybridization technique inA. victorialis var.platyphyllum. 5S rRNA genes were positioned in the intercalary regions of the short arms in homologous chromosomes 6. Two major loci of the 18S-26S rRNA genes were detected in the secondary constrictions flanking with a pair of satellite and terminal region of short arm in chromosome 4. And two additional minor loci were heterotype, representing one signal on the terminal region of the short arm in one homolog of chromsome 2, and other on one homolog of chromosome 6 with linked 5S rRNA loci. In addition chromomycin A₃ (CMA,) fluorescent banding method was used to identify the relation between Nucleolus Organizer Region (NOR) sites and CMA, positive heterochromatin sites. In homologous chromosome 4 showing 18S–26S rDNA hybridization signals revealed also distinct CMA, positive band.     

The organization of DNA in the mitotic and polytene chromosomes of Sciara coprophila

The organization of DNA in the mitotic metaphase and polytene chromosomes of the fungus gnat, Sciara coprophila, has been studied using base-specific DNA ligands, including anti-nucleoside antibodies. The DNA of metaphase and polytene chromosomes reacts with AT-specific probes (quinacrine, DAPI, Hoechst 33258 and anti-adenosine) and to a somewhat lesser extent with GC-specific probes (mithramycin, chromomycin A₃ and anticytidine). In virtually every band of the polytene chromosomes chromomycin A₃ fluorescence is almost totally quenched by counterstaining with the AT-specific ligand methyl green. This indicates that GC base pairs in most bands are closely interspersed with AT base pairs. The only exceptions are band IV-8A3 and the nucleolus organizer on the X. In contrast, quinacrine and DAPI fluorescence in every band is only slightly quenched by counterstaining with the GC-specific ligand actinomycin D. Thus, each band contains a moderate proportion of AT-rich DNA sequences with few interspersed GC base pairs. — The C-bands in mitotic and polytene chromosomes can be visualized by Giemsa staining after differential extraction of DNA and those in polytene chromosomes by the use of base-specific fluorochromes or antibodies without prior extraction of DNA. C-bands are located in the centromeric region of every chromosome, and the telomeric region of some. The C-bands in the polytene chromosomes contain AT-rich DNA sequences without closely interspered GC base pairs and lack relatively GC-rich sequences. However, one C-band in the centromeric region of chromosome IV contains relatively GC-rich sequences with closely interspersed AT base pairs. — C-bands make up less than 1% of polytene chromosomes compared to nearly 20% of mitotic metaphase chromosomes. The C-bands in polytene chromosomes are detectable with AT-specific or GC-specific probes while those in metaphase chromosomes are not. Thus, during polytenization there is selective replication of highly AT-rich and relatively GC-rich sequences and underreplication of the remainder of the DNA sequences in the constitutive heterochromatin.     

苜蓿核糖体基因物理定位及染色体荧光分带

利用核糖体基因为探针对,二倍体和四倍体苜蓿(Medicago sativa)进行原位杂交,结果表明,45s在四倍体、二倍体种中总是以单位点位于核仁组织区,5s则有2～3个位点;以二倍体种的基因组DNA为探针的原位杂交表明,蓝花苜蓿(M.coerulea)和黄花苜蓿(M.falcata)均能与四倍体染色体进行杂交,仅杂交信号强弱的染色体数目有差别;荧光染料DAPI使苜蓿的染色体显示带纹,蓝花苜蓿的DAPI带与C-带基本一致.文章对四倍体苜蓿的可能来源进行了讨论.     

Cytotaxonomic characteristics of the genus Glyptotendipes Kieffer (Chironomidae, Diptera) from fish and retention ponds (Silesia, southern Poland)

The cytotaxonomic characteristics of species of the genus Glyptotendipes (Chironomidae): G. glaucus Meigen, G. paripes Edwards, and G. barbipes (Staeger) are described. The studied material was collected from a fish pond at Go?ysz and a sewage retention pond at Chybie in Silesia. All the studied species have the chromosome set 2n = 8, but with many specific structural and functional changes. The cytogenetic data of G. glaucus showed that the studied specimens of this species have been produced by the introgressive hybridization of two sibling species: G. glaucus x G. pallens, and subsequent crossing-over in the hybrid chromosome CD. Owing to this process, the band pattern of chromosome arm D coincided with those of G. pallens. New aberrations (homo- and heterozygous deletions in arm G as well as heterozygous inversions in chromosome arm B) were detected in G. glaucus. Specific band sequences were discovered in chromosomes of G. paripes. The band patterns established in chromosomes AB and G of this species were identical with those of Siberian populations. The banding patterns of the polytene chromosomes of G. barbipes do not differ from the standard. However, high frequency of pericentric inversion of chromosome AB was established. Many new puffs were found in the polytene chromosomes of all the studied species. Their nucleolar organizer was very sensitive to environmental changes. In G. glaucus it appeared in three different states: very active, slightly active, and heterozygous state. The organic pollution existing in the sewage retention pond may contribute to possible mutations and chromosomal damage in Glypotendipes species. Structural and functional rearrangements of the polytene chromosomes of these species mobilized their genomes and provide for survival under polluted conditions.     

Homologous banding patterns in the polytene chromosomes from the larval salivary glands and ovarian nurse cells of Anopheles stephensi liston (Culicidae)

A comparison of the banding patterns of two homologous polytene chromosome arms from the larval salivary gland and ovarian nurse cell complement of Anopheles stephensi is presented. The homologous chromosomes from the somatic larval salivary glands and germ-line derived ovarian nurse cells have essentially the same band-interband organisation. An analysis of the ³H-uridine labelling patterns of a small chromosome segment from the two tissues indicates that germ-line polytene chromosomes are not radically different from somatic polytene chromosomes in their patterns of gene expression.