Rapid method to obtain high quality nuclear and chromosome images directly from living cells in the Characeae

This study focused on a method based on the capacity of cationic dyes to stain only the nucleus and chromosomes in cells subjected to either acid or alkaline hydrolysis. The method and the squash were optimized for the Characeae and were described in detail. Nuclei from vegetative shoot apices and antheridial filament cells of unfixed, fixed and herbarium material of Nitella opaca were investigated using the Azure A or Toulidine Blue stains. Comparisons with some other staining methods, used in the previous studies, were also reported. The Azure A/Toulidine Blue method is useful to obtain clear images of chromosome morphology comparable or higher to that obtained with Feulgen or Aceto‐Orcein. It requires little time and a less complicated procedure in comparison with other staining methods.     

Contribution to the genome size knowledge of New World species from the Heliantheae alliance (Asteraceae)

This article contributes first genome size assessments by flow cytometry for 16 species, 12 genera, and 3 tribes from family Asteraceae, mostly belonging to the Heliantheae alliance, an assembly of 13 tribes from subfamily Asteroideae with a large majority of its species in the New World. Most genome sizes are accompanied by their own chromosome counts, confirming in most cases, although not all, previous counts for the species, and revealing possible cases of unknown dysploidy or polyploidy for certain taxa. The data contribute to the pool of knowledge on genome size and chromosome numbers in the family Asteraceae and will further allow deeper studies and a better understanding on the role of dysploidy in the evolution of the Heliantheae alliance. However, we still lack data for tribes Chaenactideae, Neurolaeneae, Polymnieae, and Feddeeae (the latter, monospecific) to complete the alliance representation.     

黄藤染色体核型及基因组大小分析

为探究黄藤(Daemonoropsjenkinsiana)染色体核型和基因组的大小,采用体细胞染色体常规制片法与显微摄影技术相结合的方法,对黄藤染色体进行了核型分析,同时以番茄(Lycopersicon esculentum)为内标,应用流式细胞术对黄藤叶片基因组大小、DNA含量和DNA倍性进行了测定。结果表明,黄藤茎尖是理想的染色体制片材料;黄藤的染色体数为2n=24,核型公式为K(2n)=1M+17m+5sm+1st,核型类型为2C;核型不对称系数61.20%;黄藤的DNA含量为1.57 pg,基因组大小为1 539.53 Mb,黄藤的DNA倍性为二倍体(2n)。这是首次报道黄藤的核型和基因组大小,为深入开展黄藤属及其近缘属植物的核型和基因组比较分析提供了参考依据。     

Cytogeography and chromosomal variation of the endemic East Asian herb Lycoris radiata

Information on the spatial distribution of cytotypes and karyotype variation in plants is critical for studies of the origin and evolution of polyploid complexes. Here, the spatial distribution of cytological races and intraspecific variation in the karyotype of Lycoris radiata, an endemic species to East Asia, is investigated. Conventional karyotype analysis methods were used to determine ploidy level and karyotypical characteristics in 2,420 individuals from 114 populations of L. radiata nearly covering the whole distribution areas in China. Of 114 populations studied, 52 (45.61%), 58 (50.88%), and 4 (3.51%) are diploid, triploid, and mixoploid populations, respectively, with 1,224, 1,195, and 1 individuals being diploid, triploid, and tetraploid, respectively. The triploid possesses a much wider distribution range than the diploid, with the former almost occupying the entire range of this complex species in East Asia and the latter distributing in the middle and east regions of China. Triploids tend to occur at high altitudes, and the relationship between the ploidy and altitude is significantly positive but low (r² = 0.103, p < 0.01). About 98.6% of examined bulbs have a common karyotype consisting of 22 or 33 acrocentric (A) chromosomes. Some aberrant chromosomes which should be generated from A‐type chromosome have been found including metacentrics (m), small metacentrics (m′), and B‐type chromosome. The results can provide a fundamental cytogeographic data for further studies on the evolutionary origins and adaptive divergences of polyploids, especially the triploid, within L. radiata using molecular and/or ecological methods in the future.     

The presence of X- and Y-chromosomes in oocytes leads to impairment in the progression of the second meiotic division

The oocytes of B6.Y(TIR) sex-reversed female mice can be fertilized but the resultant embryos die at early cleavage stages. In the present study, we examined chromosome segregation at meiotic divisions in the oocytes of XY female mice, compared to those of XX littermates. The timing and frequency of oocyte maturation in culture were comparable between the oocytes from both types of females. At the first meiotic division, the X- and Y-chromosomes segregated independently and were retained in oocytes at equal frequencies. However, more oocytes retained the correct number of chromosomes than anticipated from random segregation. The oocytes that had reached MII-stage were activated by fertilization or incubation with SrCl(2). As expected, the majority of oocytes from XX females completed the second meiotic division and reached the 2-cell stage in 24 h. By contrast, more than half of oocytes from XY females initially remained at the MII-stage while the rest precociously entered interphase after SrCl(2) activation; very few oocytes were seen at the second anaphase or telophase and they often showed impairment of sister-chromatid separation. Eventually the majority of oocytes entered interphase and formed pronuclei, but very few reached the 2-cell stage. Similar results were obtained after fertilization. We conclude that the XY chromosomal composition in oocyte leads to impairment in the progression of the second meiotic division.     

Characterization of Japanese flounder karyotype by chromosome bandings and fluorescence in situ hybridization with DNA markers

The chromosomes of Japanese flounder, Paralichthys olivaceus, were examined by conventional differential staining methods including G-, Q-, C-, silver (Ag)-, fluorochrome, and replication R-bandings and by fluorescence in situ hybridization (FISH) with 5S and 18S rDNAs and telomeric DNA as probes. Replication R-banding substantially made it possible to identify 24 homologous pairs by their RBG-banding pattern and relative length. Both rDNA loci were mapped to chromosome 1, where 5S and 18S rDNA loci were located at the centromeric region and secondary constriction, respectively. C-banding revealed that both rDNA loci were heterochromatic, and 18S rDNA loci were positive for chromomycin A₃ but negative for 4′,6-diamidino-2-phenylindole (DAPI) staining. Telomeric FISH signals were observed at all chromosome ends and at the interstitial region of some chromosomes. The observed results were discussed in relation to the karyotype evolution in the order Pleuronectiformes.     

A simple chromosomal marker can reliably distinguishes <Emphasis Type="Italic">Poncirus</Emphasis> from <Emphasis Type="Italic">Citrus</Emphasis> species

Several chromosome types have been recognized in Citrus and related genera by chromomycin A₃ (CMA) banding patterns and fluorescent in situ hybridization (FISH). They can be used to characterize cultivars and species or as markers in hybridization and backcrossing experiments. In the present work, characterization of six cultivars of P. trifoliata (“Barnes”, “Fawcett”, “Flying Dragon”, “Pomeroy”, “Rubidoux”, “USDA”) and one P. trifoliata × C. limonia hybrid was performed by sequential analyses of CMA banding and FISH using 5S and 45S rDNA as probes. All six cultivars showed a similar CMA⁺ banding pattern with the karyotype formula 4B + 8D + 6F. The capital letters indicate chromosomal types: B, a chromosome with one telomeric and one proximal band; D, with only one telomeric band; F, without bands. In situ hybridization labeling was also similar among cultivars. Three chromosome pairs displayed a closely linked set of 5S and 45S rDNA sites, two of them co-located with the proximal band of the B type chromosomes (B/5S-45S) and the third one co-located with the terminal band of a D pair (D/5S-45S). The B/5S-45S chromosome has never been found in any citrus accessions investigated so far. Therefore, this B chromosome can be used as a marker to recognize the intergeneric Poncirus × Citrus hybrids. The intergeneric hybrid analyzed here displayed the karyotype formula 4B + 8D + 6F, with two chromosome types B/5S-45S and two D/5S-45S. The karyotype formula and the presence of two B/5S-45S chromosomes clearly indicate that the plant investigated is a symmetric hybrid. It also demonstrates the suitability of karyotype analyses to differentiate zygotic embryos or somatic cell fusions involving trifoliate orange germplasm. During the submission of this paper, we analyzed 25 other citrus cultivars with the same methodology and we found that the chromosome marker reported here can indeed distinguish Poncirus trifoliata from grapefruits, pummelos, and one variegated access of Citrus, besides the previously reported access of limes, limons, citrons, and sweet-oranges. However, among 14 mandarin cultivars, two of them displayed a single B/5S-45S chromosome, whereas in Citrus hystrix D.C., a far related species belonging to the Papeda subgenus, this chromosome type was found in homozygosis. Since these two mandarin cultivars are probably of hybrid origin, we assume that for almost all commercial cultivars and species of the subgenus Citrus this B type chromosome is a useful genetic marker.     

Polytene chromosome maps in four species of tsetse flies Glossina austeni, G. pallidipes, G. morsitans morsitans and G. m. submorsitans (Diptera: Glossinidae): a comparative analysis

Photographic polytene chromosome maps from pupal trichogen cells of four tsetse species, Glossina austeni, G. pallidipes, G. morsitans morsitans and G. m. submorsitans were constructed and compared. The homology of chromosomal elements between the species was achieved by comparing banding patterns. The telomeric and subtelomeric chromosome regions were found to be identical in all species. The pericentromeric regions were found to be similar in the X chromosome and the left arm of L1 chromosome (L1L) but different in L2 chromosome and the right arm of L1 chromosome (L1R). The L2 chromosome differs by a pericentric inversion that is fixed in the three species, G. pallidipes, G. morsitans morsitans and G. m. submorsitans. Moreover, the two morsitans subspecies appeared to be homosequential and differ only by two paracentric inversions on XL and L2L arm. Although a degree of similarity was observed across the homologous chromosomes in the four species, the relative position of specific chromosome regions was different due to chromosome inversions established during their phylogeny. However, there are regions that show no apparent homology between the species, an observation that may be attributed to the considerable intra—chromosomal rearrangements that have occurred following the species divergence. The results of this comparative analysis support the current phylogenetic relationships of the genus Glossina.     

弥漫大B细胞淋巴瘤1号染色体的基因表达分析

目的:探讨弥漫大B细胞淋巴瘤(Diffuse Large B-Cell Lymphoma,DLBCL)中1号染色体基因表达情况。方法:采用激光显微切割技术分离临床DLBCL病人淋巴结标本中的淋巴细胞,提取淋巴细胞的mRNA并与表达谱芯片杂交,通过信号扫描、处理后获得表达基因杂交信号强度。每基因设11-20对探针。杂交信号与错配探针对比,扣除背景值后,使用Wilcoxon符号秩和检验选取与错配杂交信号有显著差异的基因作为分析结果(P=0.05)。然后随机选取四个检测到的基因,使用PCR方法检验基因芯片结果的可靠性。结果:成功地从快速冷冻保存的DLBCL标本中提取RNA。使用表达谱芯片进行研究,发现了共316条1号染色体编码的基因在DLBCL细胞中表达。根据胞内定位,基因功能和基因所属的代谢通路三种分类方法对所得基因进行分类分析。基因表达密度分析显示DLBCL中1号染色体上的基因表达情况与编码基因分布情况存在统计学差异。结论:使用表达谱芯片研究了DLBCL中1号染色体上的基因表达情况。