Mitotic evidence for the tetraploid nature of Glycine max provided by high quality karyograms

The high number, very small size and morphological similarity of the chromosomes, and low metaphasic indexes obtained in root meristems have hindered the progress in cytogenetic and evolutionary studies of Glycine max. In order to contribute to the solving of these problems, we have developed a method based on the use of DNA synthesis inhibiting and anti-microtubule solutions and enzymatic maceration and air-drying techniques. Besides, we have employed a digital image analysis system tool. This method provided prometaphasic and metaphasic chromosomes showing well-defined primary and secondary constrictions, which facilitated the pairing of homologues and assembly of the first karyogram for G. max. This species possesses twenty chromosome pairs, being six metacentric and fourteen submetacentric. The karyograms support its tetraploid nature (4x = 40), specifically for the presence of chromosomes with identical morphology, and suggest that chromosome rearrangements may have occurred during the speciation of G. max.     

Homologous early replication patterns of the distal short arms of prometaphasic X and Y chromosomes

Summary Replication studies on prometaphasic human sex chromosomes reveal a distinct early replicating segment on both distal Xp and Yp. These segments correspond to high resolution bands Xp22.3 and Yp11.3. The findings demonstrate synchronous replication of these parts of the sex chromosomes and correspond to the comparatively long stretches of Xp and Yp that participate in a synaptonemal complex. Furthermore these observations are compatible with Polani's view that suggests homologous segments with similar genetic information on both sex chromosomes (Polani 1982).     

Deletion proximal to DXS68 locus (L1 probe site) in a boy with Duchenne muscular dystrophy,glycerol kinase deficiency,and adrenal hypoplasia

Summary We report a case of a boy with Duchenne muscular dystrophy (DMD) associated with GK deficiency (GK), congenital adrenal hypoplasia (AHC), and mental retardation. Cytogenetic analysis of prometaphasic chromosomes revealed an interstitial chromosome deletion at Xp21.2 possibly extending to Xp21.1 or Xp21.3. His phenotypically normal mother was heterozygous for this deletion. DNA probe analysis on Southern blots showed that the deletion affected the following probe sites: 754, pERT 84, 21A, XJ2.3, pERT 87, JBir, and J66-H1, whereas L1, C7, and CX5.4 probes gave a normal signal. Pulse field gel electrophoresis after SfiI digestion did not show abnormal fragments with L1. These data are consistent with a deletion of about 4 megabases and indicate that the GK and AHC loci are proximal to L1 and distal to J66-H1.     

The detection of the location in cattle chromosomes of a gene presumably determining testis development

The localization of the putative testis determining gene (TDF) was established with isotopic and nonisotopic methods of in situ hybridization in prometaphasic chromosomes of cattle. The results of both the methods were seen to coincide. The sites of hybridization have been revealed in X-chromosome (R-bands, q2.1 and q2.3.1). TDF was also localized in the proximal part of the long Y-chromosome arm.     

Prometaphase accumulation in murine bone marrow cells following in vivo treatment with alloxan

The aim of this study was to determine the effect of alloxan, an inhibitor of N-acetylglucosaminyl transferase that acts during the G2/M transition, on the course of mitosis in murine bone marrow cells. Mitotic cells from animals treated with different doses of alloxan were analyzed for the frequency of prometaphasic and metaphasic chromosomes based on their morphology and length. The results indicate that alloxan treatment substantially increases the frequency of prometaphase chromosomes. This suggests that N-acetylglucosaminyl transferase is also involved in the G2/M transition in bone marrow cells. Alloxan treatment also provides a method for obtaining large chromosomes for the analysis of chromosome bands, FISH and sister-chromatid exchanges.     

First Coffea arabica karyogram showing that this species is a true allotetraploid

Evolutive studies have verified that Coffea arabica (2n = 44) is a natural segmental allopolyploid originated from a cross between two diploid (2n = 22) Coffea species. Data obtained by classical cytogenetic analyses showed that C. arabica chromosomes are small and morphologically similar, which hampers the karyogram assembly with well-identified homologue pairs. In the present study, the C. arabica complement was reanalysed using an improved cytogenetic protocol that allowed the obtention of high-quality prometaphasic and metaphasic chromosomes. The results showed that chromosomes are cytogenetically distinct (1, 2, 19, 20, 21 and 22) and identical (3–4, 5–6, 7–8, 9–10, 11–12, 13–14, 15–16 and 17–18), with regard to their total length, short and long arm sizes or chromosome classes. Our work suggests that C. arabica is a true non-segmental allotetraploid but originated from different species exhibiting similar and distinct chromosomes.     

Cytologic evidence for three human X-chromosomal segments escaping inactivation

Summary Early replication of prometaphasic human sex chromosomes was studied with the bromodeoxyuridine (BrdU)-replication technique. The studies reveal that two distal segments of Xp, including bands Xp 22.13 and Xp 22.3, replicate early in S-phase and therefore may not be subject to random inactivation. Furthermore, the replication of these distal segments of Xp occurs synchronously with those of the short arm of the Y chromosome including bands Yp 11.2 and Yp 11.32. These segments of Xp and Yp correspond well to the pairing segment of the X and Y chromosomes where a synaptonemal complex forms at early pachytene of human spermatogenesis. The homologous early replication of Yp and the distal portion of Xp may be interpreted as a remnant left untouched by the differentiation of heteromorphic sex chromosomes from originally homomorphic autosomes. A third early replicating segment is situated on the long arm of the X chromosome and corresponds to band Xq 13.1. This segment may be correlated with the X-inactivation center postulated by Therman et al. (1979).     

Theoretical study of inversions affecting human chromosomes

A theoretical study of inversions affecting human chromosomes is proposed. Taking into consideration the number of bands and the fact that breaks occur at interfaces between bands, it is concluded that: 7.659 different pericentric inversions might be detected in a prometaphasic 802-band karyotype; this number decreases to 917 in a metaphasic 273-band karyotype; 8.607 and 862 different paracentric inversions might be detected in the same karyotypes respectively, but these results are likely to be overestimated. These theoretical data are used for showing that the pericentric inversions detected in human cytogenetic laboratories, are too frequently recurrent and are not distributed at random.     

High resolution R- and G-banding on the same preparation

Summary A simple method using bromodeoxyuridine (BrdU), for both cell synchronization and incorporation into replicating DNA is described. Many prophasic and prometaphasic mitoses were observed, and due to the probable blocking at different times of the cell cycle, very good R-banding and G-banding were obtained simultaneously on the same preparation.     

Chromosome elimination and sex determination in springtails (Insecta, Collembola).

A post-zygotic mechanism of sex determination is described in the two symphypleonans Dicyrtomina ornata (Nicolet) and Ptenothrix italica Dallai. The process consists of the loss of two sex chromosomes from the male embryo. At the end of the first meiotic division of spermatogenesis, a second chromosome elimination occurs, allowing half the secondary spermatocytes, later transformed into spermatids, to receive a complete haploid set of chromosomes. The secondary spermatocytes, which receive an incomplete set of chromosomes, degenerate. Males of the two collembolan species, therefore, produce a reduced number (50%) of spermatozoa. Females of D. ornata have 2n = 12 and males 2n = 10 chromosomes; females of P. italica have 2n = 14 and males 2n = 12 chromosomes. In both species, oogenesis proceeds normally and chromosomes pair and form chiasmata in meiotic prophase. The adaptive significance of this post-zygotic mechanism of sex determination is discussed. The mechanism seems to be a characteristic feature of the suborder Symphypleona. The neanurid Arthropleona Anurida maritima (Guérin), which was studied for comparative analysis, has 2n = 8 chromosomes and normal spermatogenesis producing haploid nuclei with four chromosomes. J. Exp. Zool. (Mol. Dev. Evol.) 285:215-225, 1999.     

A natural triploid in Trichomycterus davisi (Siluriformes,Trichomycteridae): mitotic and meiotic characterization by chromosome banding and synaptonemal complex analyses

Within a total of 50 analyzed specimens a male individual of Trichomycterus davisi has been recorded with 81 chromosomes including 60 metacentric, 18 submetacentric and three subtelocentric chromosomes. When compared with diploid individuals (2n = 54) and the morphological standard of chromosomes, this male is a triploid with 3n = 81 chromosomes. Since staining with silver nitrate indicates three active nucleolar organizer regions (NORs), the three NOR-bearing chromosomes in this individual are genetically active. Analysis of the synaptonemal complex (SC) by electronic microscopy shows that there is an incomplete pairing of the third set of chromosomes in the triploid individual.     

鼠颚毛厉螨染色体组型及其C-带、G-带的研究(蜱螨亚纲:厉螨科)

本文首次报道了厉螨科鼠颚毛厉螨 Tricholaelaps myonyssognathus 染色体组型及其C-带、G-带的研究。结果表明其为单二倍体性决定系统(n=6,2n=12),雄性体细胞具有6条染色体,雌性体细胞具有12条染色体。所有染色体均为单着丝粒,其着丝粒位置分别为中部(1和2号)、亚中部(3号)、亚端部(4号)及端点(5和6号)。C-带为着丝粒带。G-带的有35条深带。     

Genome scan for loci involved in cleft lip with or without cleft palate,in Chinese multiplex families

Cleft lip with or without cleft palate (CL/P) is a common congenital anomaly. Birth prevalences range from 1/500 to 1/1,000 and are consistently higher in Asian populations than in populations of European descent. Therefore, it is of interest to determine whether the CL/P etiological factors in Asian populations differ from those in white populations. A sample of 36 multiplex families were ascertained through probands with CL/P who were from Shanghai. This is the first reported genome-scan study of CL/P in any Asian population. Genotyping of Weber Screening Set 9 (387 short tandem-repeat polymorphisms with average spacing approximately 9 cM [range 1-19 cM]) was performed by the Mammalian Genotyping Service of Marshfield Laboratory. Presented here are the results for the 366 autosomal markers. Linkage between each marker and CL/P was assessed by two-point and multipoint LOD scores, as well as with multipoint heterogeneity LOD scores (HLODs) plus model-free identity-by-descent statistics and the multipoint NPL statistic. In addition, association was assessed via the transmission/disequilibrium test. LOD-score and HLOD calculations were performed under a range of models of inheritance of CL/P. The following regions had positive multipoint results (HLOD > or =1.0 and/or NPL P< or =.05): chromosomes 1 (90-110 cM), 2 (220-250 cM), 3 (130-150 cM), 4 (140-170 cM), 6 (70-100 cM), 18 (110 cM), and 21 (30-50 cM). The most significant multipoint linkage results (HLOD > or =2.0; alpha=0.37) were for chromosomes 3q and 4q. Associations with P< or =.05 were found for loci on chromosomes 3, 5-7, 9, 11, 12, 16, 20, and 21. The most significant association result (P=.009) was found with D16S769 (51 cM).     

Rapid verification of wheat-Hordeum introgressions by direct staining of SCAR, STS, and SSR amplicons.

A range of single tagged site (STS), simple sequence repeat (SSR), and sequence-characterized amplified region (SCAR) markers were screened for their utility in detecting Hordeum vulgare and H. chilense chromosomes in a wheat background. PCR conditions were optimized for specific amplification of the targeted sequences and to avoid cross-species amplification. Two H. vulgare derived STSs, six H. vulgare derived SSRs, and nine H. chilense derived SCARs were usable for the detection of five H. vulgare and three H. chilense chromosomes by direct ethidium bromide staining of the PCR products in test tubes, avoiding the more costly and time-consuming DNA electrophoresis step. The practical application of the method is illustrated by the identification of a monotelosomic substitution of H. vulgare chromosome 6HS in tritordeum and a monosomic addition of H. chilense chromosome 6Hch in durum wheat.     

Polymorphism of the Sphaerium corneum (Bivalvia, Veneroida, Sphaeriidae) revealed by cytogenetic and sequence comparison

Two populations of Sphaerium corneum were sampled from River Vilnelė and small pond in Vilnius, Lithuania. The chromosomes were studied using conventional Giemsa staining and karyometric analysis. Inter- and intra-individual variation in the diploid chromosome numbers was revealed and two different sources of chromosome variability were identified: B chromosomes and the structural changes of chromosomes of the basic (A) set. The chromosome set of the more common karyotypic form, 2 n  = 30, found in both populations, consists of all biarmed metacentric and meta-submetacentric chromosomes of gradually decreasing size. Small, biarmed, mitotically unstable B chromosomes were found in the cells of this karyotypic form. Specimens with 2 n  = 36 were found only in pond. No B chromosomes were detected in their cells. The karyotype is characterized by presence of two pairs of medium telocentrics and four pairs of small subtelocentrics. The remaining chromosomes are biarmed. Robertsonian fusions appear to be involved in formation of two karyotypic forms of S. corneum . DNA sequence analyses showed that ITS1 is identical in both karyotypic forms. On the other hand, differences in 16S sequence were revealed and two haplotypes, corresponding to two karyotypic forms, were identified. The present study opens new perspectives in establishing species-specific characters for confident identification of Sphaerium species and provides insights to the genetic intraspecific variability and possible mechanisms of speciation.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 89 , 53–64.     

The relationship between paternal age, sex ratios, and aneuploidy frequencies in human sperm, as assessed by multicolor FISH.

We studied the frequencies of X- and Y-chromosome-bearing sperm, diploidy and disomy for chromosomes 1, 12, X, and Y in sperm from 10 normal men aged 21-52 years, to determine whether there was any relationship between donor age and any of these variables. Multicolor FISH was used to control for lack of probe hybridization and to distinguish diploid sperm from disomic sperm. A minimum of 10,000 sperm per donor was evaluated for each chromosome, for a total of 225,846 sperm studied. Sperm were considered disomic if two fluorescent signals were separated by a minimal distance of one signal domain. The mean frequencies of X- and Y-bearing sperm were 50.1% and 49.0%, respectively; not significantly different from 50%. There was no correlation between paternal age and "sex ratio" in sperm. Similarly, there was no association between the frequency of diploid sperm (mean, .16%; range, .06-.42%) and donor age. For disomy frequencies, there was no relationship between donor age and disomy 12 (mean, .16%; range, .10%-.25%), XX (mean, .07%; range, .03%-.17%), and XY sperm (mean, .16%; range, .08%-.24%). There was a significant increase in the frequency of YY sperm (P = .04; mean, .18%; range, .10%-.43%) and disomy 1 sperm (P = .01; mean, .11%; range, .05%-.18%) with donor age. In summary, our results do not support a correlation between paternal age and sex ratio or diploidy.     

Chromosomal and regional localization of the genes for UMPH2, APRT, PEPD, PEPS, PSP, and PGP in mink: comparison with man and mouse

Segregation of mink biochemical markers uridine 5'-monophosphate phosphohydrolase-2 (UMPH2), adenine phosphoribosyltransferase (APRT), phosphoserine phosphatase (PSP), phosphoglycolate phosphatase (PGP), peptidases D (PEPD) and S (PEPS), as well as mink chromosomes, was investigated in a set of mink x mouse hybrid clones. The results obtained allowed us to make the following mink gene assignments: UMPH2, chromosome 8; PEPD and APRT, chromosome 7; PEPS, chromosome 6; and PSP and PGP, chromosome 14. The latter two genes are the first known markers for mink chromosome 14. For regional mapping, UMPH2 was analyzed in mouse cell clones transformed by means of mink metaphase chromosomes (Gradov et al., 1985) and also in mink x mouse hybrid clones carrying fragments of mink chromosome 8 of different sizes. Based on the data obtained, the gene for UMPH2 was assigned to the region 8pter----p26 of mink chromosome 8. The present data is compared with that previously established for man and mouse with reference to the conservation of syntenic gene groups and G-band homoeologies of chromosomes in mammals.     

Chromosome segregation from cell hybrids. VII. Reverse segregation from karyoplast hybrids suggests control by cytoplasmic factors.

Using human and Chinese hamster established lines as cell parents, we constructed hamster-human cell hybrids and human cell - hamster karyoplast hybrids. The cell hybrids retained one or two sets of hamster chromosomes and lost most of the human chromosomes. The karyoplast hybrids, however, retained a full set of human chromosomes and lost most of the Chinese hamster chromosomes. This reverse segregation pattern implies that cytoplasmic factors are major determinants of the direction of chromosome segregation.     

Study of alpha-satellite DNA in cosmid libraries, specific for chromosomes 13, 21, and 22, using fluorescence in situ hybridization

Fluorescent in situ hybridization (FISH) was employed in mapping the alpha-satellite DNA that was revealed in the cosmid libraries specific for human chromosomes 13, 21, and 22. In total, 131 clones were revealed. They contained various elements of centromeric alphoid DNA sequences of acrocentric chromosomes, including those located close to SINEs, LINEs, and classical satellite sequences. The heterochromatin of acrocentric chromosomes was shown to contain two different groups of alphoid sequences: (1) those immediately adjacent to the centromeric regions (alpha 13-1, alpha 21-1, and alpha 22-1 loci) and (2) those located in the short arm of acrocentric chromosomes (alpha 13-2, alpha 21-2, and alpha 22-2 loci). Alphoid DNA sequences from the alpha 13-2, alpha 21-2, and alpha 22-2 loci are apparently not involved in the formation of centromeres and are absent from mitotically stable marker chromosomes with a deleted short arm. Robertsonian translocations t(13q; 21q) and t(14q; 22q), and chromosome 21p-. The heterochromatic regions of chromosomes 13, 21, and 22 were also shown to contain relatively chromosome-specific repetitive sequences of various alphoid DNA families, whose numerous copies occur in other chromosomes. Pools of centromeric alphoid cosmids can be of use in further studies of the structural and functional properties of heterochromatic DNA and the identification of centromeric sequences. Moreover, these clones can be employed in high-resolution mapping and in sequencing the heterochromatic regions of the human genome. The detailed FISH analysis of numerous alphoid cosmid clones allowed the identification of several new, highly specific DNA probes of molecular cytogenetic studies--in particular, the interphase and metaphase analyses of chromosomes 2, 9, 11, 14, 15, 16, 18, 20, 21-13, 22-14, and X.