scholarly journals Establishment and Optimization of Molecular Cytogenetic Techniques (45S Rdna-FISH, GISH And Fiber-FISH) In Kiwifruit (Actinidia L.)

2021 ◽  
Author(s):  
Yang Zhao ◽  
Honghong Deng ◽  
Yao Chen ◽  
Jihan Li ◽  
Silei Chen ◽  
...  
Keyword(s):  
Fiber Quality ◽  
Molecular Cytogenetics ◽  
45S Rdna ◽  
Metaphase Chromosomes ◽  
Physical Size ◽  
Molecular Cytogenetic ◽  
Rdna Fish ◽  
Molecular Cytogenetic Techniques ◽  
First Time

Abstract Background: Kiwifruit has long been regarded as ‘the king of fruits’ for its nutritional importance. However, the molecular cytogenetics of kiwifruit has long been hampered because of the large number of basic chromosome (x=29), the inherent small size and highly similar morphology of metaphase chromosomes. Fluorescence in situ hybridization (FISH) is an indispensable molecular cytogenetic technique widely used in many plant species. Herein, the effects of post-hybridization washing temperature on FISH, blocking DNA concentration on genomic in situ hybridization (GISH), extraction method on nuclei isolation and the incubation time on the DNA fiber quality in kiwifruit were evaluated.Results: The post-hybridization washing in 2×SSC solution for 3×5 min at 37 ˚C ensured high stringency and distinct specific FISH signals in kiwifruit somatic chromosomes. The use of 50× blocking DNA provided an efficient and reliable means of discriminating between chromosomes derived from in the hybrids of A. chinensis var. chinensis (2n=2x=58) × A. eriantha Benth (2n=2x=58), and inferring the participation of parental genitors. The chopping method established in the present study were found to be very suitable for preparation of leaf nuclei in kiwifruit. A high-quality linear DNA fiber was achieved by an incubation of 20 min. The physical size of 45S rDNA signals was approximately 35-40 μmm revealed by the highly reproducible fiber-FISH procedures established and optimized in this study.Conclusions: The molecular cytogenetic techniques (45S rDNA-FISH, GISH, and high-resolution fiber-FISH) for kiwifruit was for the first time established and optimized in the present study, which is the foundation for the future genomic and evolutionary studies.

scholarly journals Application of Veterinary Cytogenetics in Domestic Animals: A Review

2019 ◽  
pp. 1-16
Author(s):  
Muhammad Sanusi Yahaya ◽  
Mohd Shahrom Salisi ◽  
Nur Mahiza Md. Isa ◽  
Abd Wahid Haron ◽  
Innocent Damudu Peter
Keyword(s):  
Chromosomal Abnormalities ◽  
Hybrid Sterility ◽  
Molecular Cytogenetics ◽  
Basic Research ◽  
Animal Production ◽  
Somatic Development ◽  
Molecular Cytogenetic ◽  
Molecular Cytogenetic Techniques

Cytogenetics is the study of chromosomes; their structure and properties, chromosome behavior during cell division, their influence on traits and factors which cause changes in chromosomes.  Veterinary cytogenetics is the application of cytogenetics to clinical problems that occur in animal production. It has been applied to understand problems such as infertility and its types, embryonic and fetal death, abnormality in sexual and somatic development and hybrid sterility and also prenatal sex determination and other forms of chromosomal abnormalities. These are achieved through conventional and banded karyotyping techniques and molecular cytogenetic techniques. Although conventional techniques are still useful and very widely applied, the nature of cytogenetics has gradually changed as a result of advances achieved in the molecular cytogenetic techniques for example fluorescent in situ hybridization and array-based techniques. These changes are evident in both molecular diagnostics and basic research. The combination of conventional and molecular cytogenetics has given rise to high resolution techniques which have enabled the study of fundamental questions regarding biological processes. It enables the study of inherited syndromes, the mechanisms of tumorigenesis at molecular level, genome organization and the determination of chromosome homologies between species. It allows the ease with which animals are selected in breeding programs and other important aspects of animal production. In this paper we discussed a number of techniques employed in cytogenetics and their methodologies, and recommend where future focus should be for the benefits of animal production.

scholarly journals Comparative molecular cytogenetic characterization of five wild Vigna species (Fabaceae)

2020 ◽  
Vol 14 (2) ◽  
pp. 243-264
Author(s):  
Chao-Wen She ◽  
Ying Mao ◽  
Xiang-Hui Jiang ◽  
Chun-Ping He
Keyword(s):  
Genomic Dna ◽  
Interspecific Variation ◽  
Rrna Genes ◽  
Fluorochrome Banding ◽  
45S Rdna ◽  
Molecular Cytogenetic ◽  
Rdna Sites ◽  
Cytogenetic Characterization ◽  
Rdna Fish

To extend our knowledge on karyotype variation of the genus Vigna Savi, 1824, the chromosomal organization of rRNA genes and fluorochrome banding patterns of five wild Vigna species were studied. Sequential combined PI (propidium iodide) and DAPI (4',6-diamidino-2-phenylindole) (CPD) staining and fluorescence in situ hybridization (FISH) with 5S and 45S rDNA probes were used to analyze the karyotypes of V. luteola (Jacquin, 1771) Bentham, 1959, V. vexillata (Linnaeus, 1753) A. Richard, 1845, V. minima (Roxburgh, 1832) Ohwi & H. Ohashi, 1969, V. trilobata (Linnaeus, 1753) Verdcourt, 1968, and V. caracalla (Linnaeus, 1753) Verdcourt,1970. For further phylogenetic analysis, genomic in situ hybridization (GISH) with the genomic DNA of V. umbellata (Thunberg, 1794) Ohwi & H.Ohashi, 1969 onto the chromosomes of five wild Vigna species was also performed. Detailed karyotypes were established for the first time using chromosome measurements, fluorochrome bands, and rDNA-FISH signals. All species had chromosome number 2n = 2x = 22, and symmetrical karyotypes that composed of only metacentric or metacentric and submetacentric chromosomes. CPD staining revealed all 45S rDNA sites in the five species analyzed, (peri)centromeric GC-rich heterochromatin in V. luteola, V. trilobata and V. caracalla, interstitial GC-rich and pericentromeric AT-rich heterochromatin in V. caracalla. rDNA-FISH revealed two 5S loci in V. caracalla and one 5S locus in the other four species; one 45S locus in V. luteola and V. caracalla, two 45S loci in V. vexillata and V. trilobata, and five 45S loci in V. minima. The karyotypes of the studied species could be clearly distinguished by the karyotypic parameters, and the patterns of the fluorochrome bands and the rDNA sites, which revealed high interspecific variation among the five species. The V. umbellata genomic DNA probe produced weak signals in all proximal regions of V. luteola and all (peri)centromeric regions of V. trilobata. The combined data demonstrate that distinct genome differentiation has occurred among the five species during evolution. The phylogenetic relationships between the five wild species and related cultivated species of Vigna are discussed based on our present and previous molecular cytogenetic data.

scholarly journals Molecular cytogenetic studies in rubber, Hevea brasiliensis Muell. Arg. (Euphorbiaceae)

Genome ◽  
1998 ◽  
Vol 41 (3) ◽  
pp. 464-467 ◽  
Author(s):  
Andrew R Leitch ◽  
K Yoong Lim ◽  
Ilia J Leitch ◽  
Michelle O'Neill ◽  
MeeLen Chye ◽  
...  
Keyword(s):  
Genetic Mapping ◽  
Ribosomal Dna ◽  
Hevea Brasiliensis ◽  
Consensus Sequence ◽  
Molecular Cytogenetics ◽  
5S Rdna ◽  
Single Pair ◽  
Interphase Nuclei ◽  
Molecular Cytogenetic

This paper reports the start of a molecular cytogenetics programme targeting the genome of the angiosperm tree species Hevea brasiliensis Muell. Arg. (rubber, 2n = 36), a major world crop about whose genetics very little is known. A metaphase karyotype of rubber is presented. In situ hybidization with the probe pTa71 for ribosomal DNA (rDNA) shows that there are four sites of probe hybidization occurring on two pairs of chromosomes called chromosomes 6 and 7 carrying sites NOR-1 and NOR-2, respectively. An examination of meristematic interphase nuclei shows that all four loci have the potential to be partially decondensed at interphase, although in many nuclei one or more loci appear fully condensed and apparently inactive. The probe pXVI revealed a single pair of chromosomes carrying 5S rDNA. The probes pTa71 and pXVI provide cytogenetic markers for three pairs of chromosomes that will be of use in genetic mapping programmes. The rubber chromosomes also have telomere sequences that hybridize with the Arabidopsis consensus sequence TTTAGGG. With the exception of the satellite region containing rDNA, which fluoresces brightly with chromomycin A3, fluorescence banding showed that there is no strong demarcation of the genome into GC- and AT-rich regions, as occurs in mammalian genomes.Key words: rubber, Hevea, genetic mapping, cytogenetics, ribosomal DNA, rDNA fluorescence banding.

scholarly journals Contributions of Cytogenetics and Molecular Cytogenetics to the Diagnosis of Adipocytic Tumors

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Jun Nishio
Keyword(s):  
Fusion Gene ◽  
Molecular Cytogenetics ◽  
Chromosomal Translocations ◽  
Molecular Cytogenetic ◽  
Mesenchymal Neoplasms ◽  
Adipocytic Tumors ◽  
Molecular Cytogenetic Techniques ◽  
Conventional Cytogenetics ◽  
Well Differentiated

Over the last 20 years, a number of tumor-specific chromosomal translocations and associated fusion genes have been identified for mesenchymal neoplasms including adipocytic tumors. The addition of molecular cytogenetic techniques, especially fluorescence in situ hybridization (FISH), has further enhanced the sensitivity and accuracy of detecting nonrandom chromosomal translocations and/or other rearrangements in adipocytic tumors. Indeed, most resent molecular cytogenetic analysis has demonstrated a translocation t(11;16)(q13;p13) that produces aC11orf95-MKL2fusion gene in chondroid lipoma. Additionally, it is well recognized that supernumerary ring and/or giant rod chromosomes are characteristic for atypical lipomatous tumor/well-differentiated liposarcoma and dedifferentiated liposarcoma, and amplification of 12q13–15 involving theMDM2,CDK4, andCPMgenes is shown by FISH in these tumors. Moreover, myxoid/round cell liposarcoma is characterized by a translocation t(12;16)(q13;p11) that fuses theDDIT3andFUSgenes. This paper provides an overview of the role of conventional cytogenetics and molecular cytogenetics in the diagnosis of adipocytic tumors.

scholarly journals Cytogenetic analyses in Trinomys (Echimyidae, Rodentia), with description of new karyotypes

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5316 ◽  
Author(s):  
Naiara Pereira Araújo ◽  
Cayo Augusto Rocha Dias ◽  
Rodolfo Stumpp ◽  
Marta Svartman
Keyword(s):  
Chromosome Mapping ◽  
Nucleolar Organizer ◽  
Interspecific Variation ◽  
Nucleolar Organizer Regions ◽  
45S Rdna ◽  
Banding Patterns ◽  
Pericentric Inversions ◽  
Cytogenetic Analyses ◽  
First Time

Trinomys Thomas (1921) is a terrestrial genus of spiny rats endemic to the Brazilian areas of Atlantic Forest and the transitional areas of Cerrado and Caatinga. Although most species have been already karyotyped, the available cytogenetic information is mostly restricted to diploid and fundamental numbers. We analyzed the chromosomes of two Trinomys species: Trinomys moojeni (2n = 56, FN = 106) and Trinomys setosus setosus (2n = 56, FN = 106 and 2n = 56, FN = 108). Our analyses included GTG- and CBG-banding, silver-staining of the nucleolar organizer regions, and chromosome mapping of telomeres and 45S rDNA by fluorescent in situ hybridization (FISH). Comparative GTG- and CBG-banding suggested that the interspecific variation may be due to rearrangements such as pericentric inversions, centromere repositioning, and heterochromatin variation. We report two new karyotypes for T. s. setosus and describe for the first time the banding patterns of the two Trinomys species.

scholarly journals Chromosome polymorphism in species of the genus Hedysarum L. (Fabaceae) originated from Southern Sibеria

2020 ◽  
Vol 19 (1) ◽  
pp. 14-16
Author(s):  
O. Yu. Yurkevich ◽  
T. E. Samatadze ◽  
I. Yu. Selyutina ◽  
S. I. Romashkina ◽  
S. A. Zoshchuk ◽  
...  
Keyword(s):  
Close Relationships ◽  
5S Rdna ◽  
Chromosomal Polymorphism ◽  
Rdna Locus ◽  
Chromosome Polymorphism ◽  
45S Rdna ◽  
High Similarity ◽  
Southern Siberia ◽  
Molecular Cytogenetic ◽  
First Time

For the first time, chromosomal polymorphism in karyotypes of three species from the section Hedysarum (= syn. Gamotion) of the genus Hedysarum L. (Fabaceae) grown in Southern Siberia has been studied with the useof molecular cytogenetic markers. This comparative molecular cytogenetic analysis revealed high similarity in morphology of chromosomes in H. alpinum L., H. austrosibiricum B. Fedtsch. and H. theinum Krasnob. as well as in patterns ofdistribution of 45S and 5S rDNA loci in their karyotypes confirming their close relationships. Considerable intra-specificpolymorphism on 45S rDNA chromosome localization was detected in H. theinum. In karyotype of H. alpinum, unlikethe other two species, two chromosome pairs bearing 5S rDNA locus were observed which could be used as additionalspecies-specific markers.

Distribution of FISH oligo-5S rDNA and oligo-(AGGGTTT)3 in Hibiscus mutabilis L.

Genome ◽  
2021 ◽  
pp. 1-10
Author(s):  
Xiaomei Luo ◽  
Zhoujian He
Keyword(s):  
Chromosome Number ◽  
Physical Map ◽  
Molecular Cytogenetics ◽  
5S Rdna ◽  
Chromosome Length ◽  
Chromosome Identification ◽  
Karyotype Asymmetry ◽  
First Time ◽  
Distribution Of Fish

Hibiscus exhibits high variation in chromosome number both within and among species. The Hibiscus mutabilis L. karyotype was analyzed in detail using fluorescence in situ hybridization (FISH) with oligonucleotide probes for (AG3T3)3 and 5S rDNA, which were tested here for the first time. In total, 90 chromosomes were counted in prometaphase and metaphase, and all exhibited similarly intense (AG3T3)3 signals at both ends. (AG3T3)3 showed little variation and thus did not allow discrimination among H. mutabilis chromosomes, but its location at both ends confirmed the integrity of each chromosome, thus contributing to accurate counting of the numerous, small chromosomes. Oligo-5S rDNA marked the proximal/distal regions of six chromosomes: weak signals on chromosomes 7 and 8, slightly stronger signals on chromosomes 15 and 16, and very strong signals on chromosomes 17 and 18. Therefore, 5S rDNA could assist in chromosome identification in H. mutabilis. Metaphase chromosome lengths ranged from 3.00 to 1.18 μm, indicating small chromosomes. The ratios of longest to shortest chromosome length in prometaphase and metaphase were 2.58 and 2.54, respectively, indicating karyotype asymmetry in H. mutabilis. These results provide an exact chromosome number and a physical map, which will be useful for genome assembly and contribute to molecular cytogenetics in the genus Hibiscus.

Use of meiotic FISH for identification of a new monosome in Gossypium hirsutum L.

Genome ◽  
1997 ◽  
Vol 40 (1) ◽  
pp. 34-40 ◽  
Author(s):  
Yuanfu Ji ◽  
Dwaine A. Raska ◽  
M. Nurul Islam-Faridi ◽  
Charles F. Crane ◽  
Michael S. Zwick ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Gossypium Hirsutum ◽  
Fluorescence In Situ Hybridization ◽  
Molecular Cytogenetics ◽  
Clinical Diagnostics ◽  
Identification System ◽  
Gossypium Hirsutum L ◽  
Molecular Cytogenetic ◽  
Cytogenetic Methods

The extensive use of molecular cytogenetics in human genetics and clinical diagnostics indicates that analogous applications in plants are highly feasible. One sort of application would be the identification of new aneuploids, which traditionally involves either direct karyotypic identification, which is feasible in only a few plant species, or tests with markers (cytogenetic, genetic, or molecular), which require sexual hybridization and at least one subsequent seed or plant generation. We have used meiotic fluorescence in situ hybridization (FISH) to analyze a new monosome of cotton (Gossypium hirsutum L., 2n = 4x = 52, 2(AD)1) that had a phenotype which seemed to be distinct from monosomes in the Cotton Cytogenetic Collection. Painting with A2-genome DNA revealed the monosome's D-subgenome origin. DAPI–PI staining showed that the monosome carries a major NOR, delimiting it to the major NOR-bearing chromosomes of the D-subgenome, i.e., 16 or 23. Dual-color FISH with 5S and 18S–28S rDNAs indicated that the monosome contains separate major clusters of each of these two tandemly repeated rDNA elements, thus delimiting the monosome to chromosome 23, for which the Cotton Cytogenetic Collection has previously been devoid of any sort of deficiency. Of the 26 chromosomes in the cotton genome, the Collection now provides coverage for 16 (70%) in the form of monosomy, and 20 (77%) in the form of monosomy and (or) telosomy. Use of molecular cytogenetic methods to identify a new plant aneuploid in cotton exemplifies the fact that a physicochemical karyotypic chromosome identification system is not required a priori for application of new molecular cytogenetic methods, thus indicating their potential applicability to nearly all plant species.Key words: fluorescence in situ hybridization, monosome, aneuploid, Gossypium hirsutum.

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.

scholarly journals Genotype-Phenotype Characterization of Wolf-Hirschhorn Syndrome Confirmed by FISH: Case Reports

2012 ◽  
Vol 2012 ◽  
pp. 1-5 ◽  
Author(s):  
F. Sheth ◽  
O. R. Akinde ◽  
C. Datar ◽  
O. V. Adeteye ◽  
J. Sheth
Keyword(s):  
Cytogenetic Study ◽  
Case Reports ◽  
Normal Karyotype ◽  
Comparative Genomic ◽  
Molecular Cytogenetic ◽  
Contiguous Gene ◽  
Multiple Malformation ◽  
Molecular Cytogenetic Techniques

The Wolf-Hirschhorn syndrome (WHS) is a multiple malformation and contiguous gene syndrome resulting from the deletion encompassing a 4p16.3 region. A microscopically visible terminal deletion on chromosome 4p (4p16→pter) was detected in Case 1 with full blown features of WHS. The second case which had an interstitial microdeletion encompassingWHSC 1andWHSC 2genes at 4p16.3 presented with less striking clinical features of WHS and had an apparently “normal” karyotype. The severity of the clinical presentation was as a result of haploinsufficiency and interaction with surrounding genes as well as mutations in modifier genes located outside the WHSCR regions. The study emphasized that an individual with a strong clinical suspicion of chromosomal abnormality and a normal conventional cytogenetic study should be further investigated using molecular cytogenetic techniques such as fluorescencein situhybridization (FISH) or array-comparative genomic hybridization (a-CGH).

Sign in / Sign up

Export Citation Format

Share Document