Molecular cytogenetics of Icelandic birch species: physical mapping by in situ hybridization and rDNA polymorphism

1995 ◽  
Vol 25 (1) ◽  
pp. 101-108 ◽  
Author(s):  
Kesara Anamthawat-Jonsson ◽  
J.S. Heslop-Harrison
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
Molecular Cytogenetics ◽  
Ribosomal Gene ◽  
Nucleolus Organizer ◽  
Environmental Conservation ◽  
Nucleolus Organizer Regions ◽  
A Genome ◽  
Breeding Programmes

The physical mapping of genes can reveal the organization of a genome and identify relationships of plant species, especially where they are involved in interspecific hybridization and polyploidy. Here we determine the chromosomal locations of the major ribosomal gene family (18S-5.8S-26S rDNA) by fluorescent in situ hybridization in two Icelandic birch species, Betulapubescens Ehrh. and Betulanana L. In the tetraploid birch (B. pubescens), the rDNA was localized on four major and two minor sites, while the diploid dwarf birch (B. nana) had four major sites. The major loci in both species were in nucleolus organizer regions, close to the centromeres of a pair of metacentric and a pair of sub-metacentric chromosomes. The dispersed interphase in situ hybridization pattern showed gene expression at all major sites. The two additional loci in B. pubescens, when detected, appeared to be sub-telomeric and inactive at interphase. Southern analysis of rDNA showed considerable restriction fragment length polymorphism in B. pubescens. Some polymorphism may reflect gene flow among populations and between the two co-existing birch species. The understanding of genome relationships, gene introgression, and evolution of birch species will be important to the breeding programmes steered towards environmental conservation and forestry.

scholarly journals Physical mapping of repetitive oligonucleotides facilitates the establishment of a genome map-based karyotype to identify chromosomal variations in peanut

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Liuyang Fu ◽  
Qian Wang ◽  
Lina Li ◽  
Tao Lang ◽  
Junjia Guo ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
Tandem Repeats ◽  
Genetic Research ◽  
Diploid Species ◽  
Reference Sequence ◽  
A Genome ◽  
Genome Map ◽  
Chromosomal Variants

Abstract Background Chromosomal variants play important roles in crop breeding and genetic research. The development of single-stranded oligonucleotide (oligo) probes simplifies the process of fluorescence in situ hybridization (FISH) and facilitates chromosomal identification in many species. Genome sequencing provides rich resources for the development of oligo probes. However, little progress has been made in peanut due to the lack of efficient chromosomal markers. Until now, the identification of chromosomal variants in peanut has remained a challenge. Results A total of 114 new oligo probes were developed based on the genome-wide tandem repeats (TRs) identified from the reference sequences of the peanut variety Tifrunner (AABB, 2n = 4x = 40) and the diploid species Arachis ipaensis (BB, 2n = 2x = 20). These oligo probes were classified into 28 types based on their positions and overlapping signals in chromosomes. For each type, a representative oligo was selected and modified with green fluorescein 6-carboxyfluorescein (FAM) or red fluorescein 6-carboxytetramethylrhodamine (TAMRA). Two cocktails, Multiplex #3 and Multiplex #4, were developed by pooling the fluorophore conjugated probes. Multiplex #3 included FAM-modified oligo TIF-439, oligo TIF-185-1, oligo TIF-134-3 and oligo TIF-165. Multiplex #4 included TAMRA-modified oligo Ipa-1162, oligo Ipa-1137, oligo DP-1 and oligo DP-5. Each cocktail enabled the establishment of a genome map-based karyotype after sequential FISH/genomic in situ hybridization (GISH) and in silico mapping. Furthermore, we identified 14 chromosomal variants of the peanut induced by radiation exposure. A total of 28 representative probes were further chromosomally mapped onto the new karyotype. Among the probes, eight were mapped in the secondary constrictions, intercalary and terminal regions; four were B genome-specific; one was chromosome-specific; and the remaining 15 were extensively mapped in the pericentric regions of the chromosomes. Conclusions The development of new oligo probes provides an effective set of tools which can be used to distinguish the various chromosomes of the peanut. Physical mapping by FISH reveals the genomic organization of repetitive oligos in peanut chromosomes. A genome map-based karyotype was established and used for the identification of chromosome variations in peanut following comparisons with their reference sequence positions.

Genetic and cytogenetic analyses of the A genome of Triticum monococcum. VI. Production and identification of primary trisomics using the C-banding technique

Genome ◽  
1990 ◽  
Vol 33 (4) ◽  
pp. 542-555 ◽  
Author(s):  
B. Friebe ◽  
N.-S. Kim ◽  
J. Kuspira ◽  
B. S. Gill
Keyword(s):  
Nucleolus Organizer ◽  
Triticum Monococcum ◽  
Banding Patterns ◽  
Primary Trisomics ◽  
Nucleolus Organizer Regions ◽  
C Banding ◽  
A Genome ◽  
Chromosome 5A ◽  
Triple Dose

Cytogenetic studies in Triticum monococcum (2n = 2x = 14) are nonexistent. To initiate such investigations in this species, a series of primary trisomics was generated from autotriploids derived from crosses between induced autotetraploids and diploids. All trisomics differed phenotypically from their diploid progenitors. Only two of the seven possible primary trisomic types produced distinct morphological features on the basis of which they could be distinguished. The chromosomes in the karyotype were morphologically very similar and could not be unequivocally identified using standard techniques. Therefore, C-banding was used to identify the chromosomes and trisomics of this species. Ag–NOR staining and in situ hybridization, using rDNA probes, were used to substantiate these identifications. A comparison of the C-banding patterns of the chromosomes of T. monococcum with those of the A genome in Triticum aestivum permitted identification of five of its chromosomes, viz., 1A, 2A, 3A, 5A, and 7A. The two remaining chromosomes possessed C-banding patterns that were not equivalent to those of any of the chromosomes in the A genome of the polyploid wheats. When one of these undesignated chromosomes from T. monococcum var. boeoticum was substituted for chromosome 4A of Triticum turgidum, it compensated well phenotypically and therefore genetically for the loss of this chromosome in the recipient species. Because this T. monococcum chromosome appeared to be homoeologous to the group 4 chromosomes of polyploid wheats, it was designated 4A. By the process of elimination the second undesignated chromosome in T. monococcum must be 6A. Analysis of the trisomics obtained led to the following conclusions. (i) Trisomics for chromosome 3A were not found among the trisomic lines analyzed cytologically. (ii) Primary trisomics for chromosomes 2A, 4A, 6A, and 7A were positively identified. (iii) Trisomics for the SAT chromosomes 1A and 5A were positively identified in some cases and not in others because of polymorphism in the telomeric C-band of the short arm of chromosome 1A. (iv) Trisomics for chromosome 7A were identified on the basis of their distinct phenotype, viz., the small narrow heads and small narrow leaves. Because rRNA hybridizes lightly to nucleolus organizer regions on chromosome 1A and heavily to nucleolus organizer regions on chromosome 5A, our results indicate that trisomics in line 50 carry chromosome 1A in triple dose and trisomics in lines 28 and 51 carry chromosome 5A in triplicate. Variable hybridization of the rDNA probe to nucleolus organizer regions on chromosomes in triple dose in lines 7, 20, and 28 precluded the identification of the extra chromosome in these lines. Cytogenetic methods for unequivocally identifying trisomics for chromosomes 1A and 5A are discussed. Thus six of the series of primary trisomics have been identified. Telotrisomic lines are also being produced.Key words: Triticum monococcum, trisomics, C-banding, Ag-NOR staining, in situ hybridization, rDNA probes, plant morphology.

Nonisotopic in situ hybridization and plant genome mapping: the first 10 years

Genome ◽  
1994 ◽  
Vol 37 (5) ◽  
pp. 717-725 ◽  
Author(s):  
Jiming Jiang ◽  
Bikram S. Gill
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
Dna Sequences ◽  
Genome Mapping ◽  
Molecular Cytogenetics ◽  
Gene Families ◽  
Single Copy ◽  
Plant Genome ◽  
Metaphase Chromosomes

Nonisotopic in situ hybridization (ISH) was introduced in plants in 1985. Since then the technique has been widely used in various areas of plant genome mapping. ISH has become a routine method for physical mapping of repetitive DNA sequences and multicopy gene families. ISH patterns on somatic metaphase chromosomes using tandemly repeated sequences provide excellent physical markers for chromosome identification. Detection of low or single copy sequences were also reported. Genomic in situ hybridization (GISH) was successfully used to analyze the chromosome structure and evolution of allopolyploid species. GISH also provides a powerful technique for monitoring chromatin introgession during interspecific hybridization. A sequential chromosome banding and ISH technique was developed. The sequential technique is very useful for more precise and efficient mapping as well as cytogenetic determination of genomic affinities of individual chromosomes in allopolyploid species. A critical review is made on the present resolution of the ISH technique and the future outlook of ISH research is discussed.Key words: in situ hybridization, physical mapping, genome mapping, molecular cytogenetics.

Karyotyping Melandrium album, a dioecious plant with heteromorphic sex chromosomes

Genome ◽  
1990 ◽  
Vol 33 (4) ◽  
pp. 556-562 ◽  
Author(s):  
D. D. Ciupercescu ◽  
J. Veuskens ◽  
A. Mouras ◽  
D. Ye ◽  
M. Briquet ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Sex Chromosomes ◽  
Nucleolus Organizer ◽  
Rrna Genes ◽  
Hybridization Technique ◽  
Metaphase Chromosomes ◽  
Nucleolus Organizer Regions ◽  
Melandrium Album ◽  
Group B

Mitotic metaphase chromosomes of Melandrium album obtained from root protoplasts were studied. Morphologically, the chromosomes were either metacentrics or submetacentrics. They were classified into three distinct groups: group A comprising six pairs of autosomal metacentrics, group B comprising five pairs of autosomal submetacentrics, and the sex chromosomes: X and Y. The X chromosome is a metacentric (r = 1.44), which accounts for more than 14% of the genome. The Y chromosome is a metacentric with, virtually, equal arms (r = 1.09) and accounts for 21% of the genome, being the largest of the complement. The Y:X ratio was 1.4. Ethidium bromide, caffeine, and vinblastine were used to obtain a better resolution and higher frequency of satellited chromosomes 7q and 9p. The proposed karyotype of M. album is 2n = 24, XX, s(7q;9p) for female and 2n = 24, XY, s(7q;9p) for male plants. Nucleolus organizer regions (NORs) were present at the telomeric sites of three chromosome pairs: 7q, 9p, and 10p. The NORs were polymorphic, particularly between the nonhomologous chromosomes. The in situ hybridization technique localized the rRNA genes on four chromosome pairs: 5p, 7q, 9p, and 10p. The discrepancy between the NORs and the hybridization signals was probably due to the fact that NORs were restricted only to transcriptionally active rRNA genes. It was concluded that for a complete description and characterization of rRNA genes, both NOR detection and in situ hybridization techniques, as complementary methods, should be employed.Key words: Melandrium album, karyotype, satellites, idiogram, nucleolus organizer regions, in situ hybridization.

scholarly journals Karyotype variability in neotropical catfishes of the family Pimelodidae (Teleostei: Siluriformes)

2011 ◽  
Vol 9 (1) ◽  
pp. 97-105 ◽  
Author(s):  
Américo Moraes Neto ◽  
Maelin da Silva ◽  
Daniele Aparecida Matoso ◽  
Marcelo Ricardo Vicari ◽  
Mara Cristina de Almeida ◽  
...  
Keyword(s):  
Fluorescent In Situ Hybridization ◽  
Diploid Number ◽  
Molecular Cytogenetics ◽  
Ribosomal Genes ◽  
Nucleolus Organizer ◽  
Nucleolus Organizer Regions ◽  
Karyotype Variability ◽  
Interspecific Variations ◽  
The Family

Karyotypic data are presented for four species of fish belonging to the Pimelodidae family. These species show a conserved diploid number, 2n = 56 chromosomes, with different karyotypic formulae. The analyzed species showed little amount of heterochromatin located preferentially in the centromeric and telomeric regions of some chromosomes. The nucleolus organizer regions activity (Ag-NORs) and the chromosomal location of ribosomal genes by fluorescent in situ hybridization (FISH), with 18S and 5S probes, showing only one chromosome pair marked bearer of ribosomal genes, the only exception was Pimelodus britskii that presented multiple NORs and syntenic location of the 18S and 5S probes. Non-Robertsonian events, as pericentric inversion and NORs duplication are requested to explain the karyotype diversification in Pseudoplatystoma from the rio Paraguay (MS), Pimelodus from the rio Iguaçu (PR), Sorubim from the rio Paraguay (MS) and Steindachneridion from the rio Paraíba do Sul (SP). The obtained data for the karyotype macrostructure of these species corroborates a conserved pattern observed in Pimelodidae. On the other hand, interspecific variations detected by molecular cytogenetics markers made possible cytotaxonomic inferences and differentiation of the species here analyzed.

An efficient method for the physical mapping of transgenes in barley using in situ hybridization

Genome ◽  
2001 ◽  
Vol 44 (1) ◽  
pp. 104-110 ◽  
Author(s):  
H Salvo-Garrido ◽  
S Travella ◽  
T Schwarzacher ◽  
W A Harwood ◽  
J W Snape
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
Efficient Method ◽  
Probe Design ◽  
Nick Translation ◽  
Breeding Programmes ◽  
Detection Probe ◽  
Translation Mixture ◽  
Transgene Detection

The genetic transformation of crops by particle bombardment and Agrobacterium tumefaciens systems have the potential to complement conventional plant breeding programmes. However, before deployment, transgenic plants need to be characterized in detail, and physical mapping is an integral part of this process. Therefore, it is important to have a highly efficient method for transgene detection by fluorescence in situ hybridization (FISH). This study describes a new approach, which provides efficient control of probe length and labelling, both of which play an important role in in situ hybridization of transgenes. The approach is based on reducing the size of the plasmid prior to labelling by nick translation, rather than using the whole or linearized plasmid, or varying the amounts of DNaseI in the nick translation mixture. This provided much more efficient labelling of the probe, which yielded optimal hybridization, minimal fluorescent background, and accurate physical location of the transgene.Key words: barley, transformation, FISH, transgene detection, probe design.

scholarly journals Cytogenetic Characterization of the Genome of Interspecies Hybrids (Alopex-Vulpes)

2015 ◽  
Vol 15 (1) ◽  
pp. 81-91 ◽  
Author(s):  
Monika Bugno-Poniewierska ◽  
Klaudia Pawlina ◽  
Natalia Orszulak-Wolny ◽  
Bartosz Woźniak ◽  
Maciej Wnuk ◽  
...  
Keyword(s):  
Interspecific Hybrids ◽  
Molecular Cytogenetics ◽  
Nucleolus Organizer ◽  
Red Foxes ◽  
Nucleolus Organizer Regions ◽  
Arctic Foxes ◽  
Interspecies Hybrids ◽  
A Genome ◽  
Metaphase Stage ◽  
Cytogenetic Characterization

AbstractCreation of interspecific hybrids is widely common among plants and animals in order to improve economically important traits for humans. The studied material consisted of chromosomal preparations in the metaphase stage obtained from interspecies hybrids of arctic foxes (Alopex lagopus) and red foxes (Vulpes vulpes). The aim of the study was to analyze the karyotype of the interspecific hybrids taking into account the number of chromosomes of sets A and B. With the use of techniques of classical cytogenetics (C bands, AgNOR bands) and molecular cytogenetics (FISH, PRINS) we carried out a genome analysis of Alopex-Vulpes hybrids. The results of this study showed that chromosomal markers of the interspecies hybrids are inherited from the parent species and are a result of combination of their two genomes. However, intraindividual differences are also observed which may result from aberrations of chromosome segregation during embryonic development. This may lead to the formation of different cell lines with different karyotypes (mosaicism). Moreover, chromosomes of the interspecies hybrids showed telomeric signals at the ends, in the centromers, as well as short chromosome arm rich in heterochromatin. The use of PRINS method led to identification of nucleolus organizer regions on 12 chromosomes of the interspecies hybrids. The hybridization signals obtained were characterized by different size and intensity. In addition, single copies of rDNA in the centromeric regions of several metacentric chromosomes were identified.

Chromosomal organization of a sequence related to LTR-like elements of Ty1-copia retrotransposons in Avena species

Genome ◽  
1999 ◽  
Vol 42 (4) ◽  
pp. 706-713 ◽  
Author(s):  
Concha Linares ◽  
Antonio Serna ◽  
Araceli Fominaya
Keyword(s):  
In Situ Hybridization ◽  
Diploid Species ◽  
D Genome ◽  
Specific Sequence ◽  
Chromosomal Organization ◽  
Intergenomic Translocations ◽  
A Genome ◽  
Slot Blot Analysis ◽  
Copia Retrotransposons

A repetitive sequence, pAs17, was isolated from Avena strigosa (As genome) and characterized. The insert was 646 bp in length and showed 54% AT content. Databank searches revealed its high homology to the long terminal repeat (LTR) sequences of the specific family of Ty1-copia retrotransposons represented by WIS2-1A and Bare. It was also found to be 70% identical to the LTR domain of the WIS2-1A retroelement of wheat and 67% identical to the Bare-1 retroelement of barley. Southern hybridizations of pAs17 to diploid (A or C genomes), tetraploid (AC genomes), and hexaploid (ACD genomes) oat species revealed that it was absent in the C diploid species. Slot-blot analysis suggested that both diploid and tetraploid oat species contained 1.3 × 104 copies, indicating that they are a component of the A-genome chromosomes. The hexaploid species contained 2.4 × 104 copies, indicating that they are a component of both A- and D-genome chromosomes. This was confirmed by fluorescent in situ hybridization analyses using pAs17, two ribosomal sequences, and a C-genome specific sequence as probes. Further, the chromosomes involved in three C-A and three C-D intergenomic translocations in Avena murphyi (AC genomes) and Avena sativa cv. Extra Klock (ACD genomes), respectively, were identified. Based on its physical distribution and Southern hybridization patterns, a parental retrotransposon represented by pAs17 appears to have been active at least once during the evolution of the A genome in species of the Avena genus.Key words: chromosomal organization, in situ hybridization, intergenomic translocations, LTR sequence, oats.

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