FISH-mapping of rDNAs and Arabidopsis BACs on pachytene complements of selected Brassicas

Genome ◽  
2002 ◽  
Vol 45 (1) ◽  
pp. 189-197 ◽  
Author(s):  
Piotr A Ziolkowski ◽  
Jan Sadowski
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
5S Rdna ◽  
Artificial Chromosome ◽  
Rdna Locus ◽  
Fish Analysis ◽  
45S Rdna ◽  
Fish Mapping ◽  
Rdna Sequences

To improve resolution of physical mapping on Brassica chromosomes, we have chosen the pachytene stage of meiosis where incompletely condensed bivalents are much longer than their counterparts at mitotic metaphase. Mapping with 5S and 45S rDNA sequences demonstrated the advantage of pachytene chromosomes in efficient physical mapping and confirmed the presence of a novel 5S rDNA locus in Brassica oleracea, initially identified by genetic mapping using restriction fragment length polymorphism (RFLP). Fluorescence in situ hybridization (FISH) analysis visualized the presence of the third 5S rDNA locus on the long arm of chromosome C2 and confirmed the earlier reports of two 45S rDNA loci in the B. oleracea genome. FISH mapping of low-copy sequences from the Arabidopsis thaliana bacterial artificial chromosome (BAC) clones on the B. oleracea chromosomes confirmed the expectation of efficient and precise physical mapping of meiotic bivalents based on data available from A. thaliana and indicated conserved organization of these two BAC sequences on two B. oleracea chromosomes. Based on the heterologous in situ hybridization with BACs and their mapping applied to long pachytene bivalents, a new approach in comparative analysis of Brassica and A. thaliana genomes is discussed.Key words: Brassicaceae, pachytene chromosomes, FISH, rDNA, BACs.

Physical mapping of four sites of 5S rDNA sequences and one site of the α-amylase-2 gene in barley (Hordeum vulgare)

Genome ◽  
1993 ◽  
Vol 36 (3) ◽  
pp. 517-523 ◽  
Author(s):  
I. J. Leitch ◽  
J. S. Heslop-Harrison
Keyword(s):  
In Situ Hybridization ◽  
Hordeum Vulgare ◽  
Physical Mapping ◽  
5S Rdna ◽  
Chromosome 1 ◽  
Chromosome 2 ◽  
Hordeum Vulgare L ◽  
Rdna Sequences ◽  
5S Rdna Sequence

The 5S rDNA sequences have been mapped on four pairs of barley (Hordeum vulgare L.) chromosomes using in situ hybridization and barley monotelotrisomic lines. The 5S rDNA sequences are located, genetically and physically, on the short arm of chromosome 1 (7I) and the long arms of chromosomes 2 (2I) and 3 (3I). The 5S rDNA sequence is also located on the physically long arm of chromosome 4 (4I). Only one site on chromosome 2(2I) has previously been reported. The characteristic locations of the 5S rDNA sequences make them useful as molecular markers to identify each barley chromosome. The physical position of the low-copy α-amylase-2 gene was determined using in situ hybridization; the location of this gene on the long arm of chromosome 1 (7I) was confirmed by reprobing the same preparation with the 5S rDNA probe. The results show that there is a discrepancy between the physical and genetic position of the α-amylase-2 gene.Key words: genetic mapping, physical mapping, barley, mapping, 5S DNA, α-amylase, in situ hybridization.

Molecular cytogenetic analysis ofAegilops cylindrica Host

Genome ◽  
1999 ◽  
Vol 42 (3) ◽  
pp. 497-503 ◽  
Author(s):  
Gabriella Linc ◽  
Bernd R Friebe ◽  
Ralf G Kynast ◽  
Marta Molnar-Lang ◽  
Bela Köszegi ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
5S Rdna ◽  
Rdna Locus ◽  
Fish Analysis ◽  
Aegilops Cylindrica ◽  
Fish Genome ◽  
C Banding ◽  
Rdna Loci ◽  
25S Rdna

The genomic constitution of Aegilops cylindrica Host (2n = 4x = 28, DcDcCcCc) was analyzed by C-banding, genomic in situ hybridization (GISH), and fluorescence in situ hybridization (FISH) using the DNA clones pSc119, pAs1, pTa71, and pTA794. The C-banding patterns of the Dc- and Cc-genome chromosomes of Ae. cylindrica are similar to those of D-and C-genome chromosomes of the diploid progenitor species Ae. tauschii Coss. and Ae. caudata L., respectively. These similarities permitted the genome allocation and identification of the homoeologous relationships of the Ae. cylindrica chromosomes. FISH analysis detected one major 18S-5.8S-25S rDNA locus in the short arm of chromosome 1Cc. Minor 18S-5.8S-25S rDNA loci were mapped in the short arms of 5Dc and 5Cc. 5S rDNA loci were identified in the short arm of chromosomes 1Cc, 5Dc, 5Cc, and 1Dc. GISH analysis detected intergenomic translocation in three of the five Ae. cylindrica accessions. The breakpoints in all translocations were non-centromeric with similar-sized segment exchanges.Key words: Aegilops cylindrica, C-banding, GISH, FISH, genome evolution.

scholarly journals Fluorescence In Situ Hybridization (FISH) Analysis of the Locations of the Oligonucleotides 5S rDNA, (AGGGTTT)3, and (TTG)6 in Three Genera of Oleaceae and Their Phylogenetic Framework

Genes ◽  
2019 ◽  
Vol 10 (5) ◽  
pp. 375 ◽  
Author(s):  
Xiaomei Luo ◽  
Juncheng Liu
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
5S Rdna ◽  
Fish Analysis ◽  
Cytogenetic Map ◽  
Ligustrum Lucidum ◽  
Starting Point ◽  
Central Regions ◽  
Almost All

We report the cytogenetic map for a collection of species in the Oleaceae, and test similarities among the karyotypes relative to their known species phylogeny. The oligonucleotides 5S ribosomal DNA (rDNA), (AGGGTTT)3, and (TTG)6 were used as fluorescence in situ hybridization (FISH) probes to locate the corresponding chromosomes in three Oleaceae genera: Fraxinus pennsylvanica, Syringa oblata, Ligustrum lucidum, and Ligustrum × vicaryi. Forty-six small chromosomes were identified in four species. (AGGGTTT)3 signals were observed on almost all chromosome ends of four species, but (AGGGTTT)3 played no role in distinguishing the chromosomes but displayed intact chromosomes and could thus be used as a guide for finding chromosome counts. (TTG)6 and 5S rDNA signals discerned several chromosomes located at subterminal or central regions. Based on the similarity of the signal pattern (mainly in number and location and less in intensity) of the four species, the variations in the 5S rDNA and (TTG)6 distribution can be ordered as L. lucidum < L. × vicaryi < F. pennsylvanica < S. oblata. Variations have observed in the three genera. The molecular cytogenetic data presented here might serve as a starting point for further larger-scale elucidation of the structure of the Oleaceae genome, and comparison with the known phylogeny of Oleaceae family.

Comparative physical mapping of the 18S-5.8S-26S rDNA in three sorghum species

Genome ◽  
2000 ◽  
Vol 43 (5) ◽  
pp. 918-922 ◽  
Author(s):  
Yijun Sang ◽  
George H Liang
Keyword(s):  
In Situ Hybridization ◽  
Sorghum Bicolor ◽  
Fluorescence In Situ Hybridization ◽  
Chromosome Evolution ◽  
Rdna Locus ◽  
Sorghum Halepense ◽  
Terminal Position ◽  
26S Rdna ◽  
Rdna Sequences

The physical locations of the 18S-5.8S-26S rDNA sequences were examined in three sorghum species by fluorescence in situ hybridization (FISH) using biotin-labeled heterologous 18S-5.8S-26S rDNA probe (pTa71). Each 18S-5.8S-26S rDNA locus occurred at two sites on the chromosomes in Sorghum bicolor (2n = 20) and S. versicolor (2n = 10), but at four sites on the chromosomes of S. halepense (2n = 40) and the tetraploid S. versicolor (2n = 20). Positions of the rDNA loci varied from the interstitial to terminal position among the four accessions of the three sorghum species. The rDNA data are useful for investigation of chromosome evolution and phylogeny. This study excluded S. versicolor as the possible progenitor of S. bicolor.Key words: Sorghum bicolor, Sorghum versicolor, Sorghum halepense, 18S-5.8S-26S rDNA, fluorescence in situ hybridization.

Genetic and chromosomal localization of the 5S rDNA locus in sugar beet (Beta vulgaris L.)

Genome ◽  
1997 ◽  
Vol 40 (2) ◽  
pp. 171-175 ◽  
Author(s):  
J. Schondelmaier ◽  
T. Schmidt ◽  
C. Jung ◽  
J. S. Heslop-Harrison
Keyword(s):  
In Situ Hybridization ◽  
Linkage Group ◽  
Sugar Beet ◽  
Beta Vulgaris ◽  
5S Rdna ◽  
Rdna Locus ◽  
5S Rrna ◽  
Rrna Genes ◽  
Rrna Gene

A digoxigenin-labelled 5S rDNA probe containing the 5S rRNA gene and the adjacent intergenic spacer was used for in situ hybridization to metaphase and interphase chromosomes of a trisomic stock from sugar beet (Beta vulgaris L.). Three chromosomes of primary trisomic line IV (T. Butterfass. Z. Bot. 52: 46–77. 1964) revealed signals close to the centromeres. Polymorphisms of 5S rDNA repeats in a segregating population were used to map genetically the 5S rRNA genes within a cluster of markers in linkage group II of sugar beet. The concentration of genetic markers around the centromere presumably reflects the suppressed recombination frequency in centromeric regions. The correlation of physical and genetic data allowed the assignment of a linkage group to sugar beet chromosome IV according to line IV of the primary trisomics.Key words: Beta vulgaris, sugar beet, 5S rRNA, in situ hybridization, RFLPs, trisomics.

Karyotype analysis of Lilium longiflorum and Lilium rubellum by chromosome banding and fluorescence in situ hybridisation

Genome ◽  
2001 ◽  
Vol 44 (5) ◽  
pp. 911-918 ◽  
Author(s):  
Ki-Byung Lim ◽  
Jannie Wennekes ◽  
J Hans de Jong ◽  
Evert Jacobsen ◽  
Jaap M van Tuyl
Keyword(s):  
In Situ Hybridisation ◽  
5S Rdna ◽  
Lilium Longiflorum ◽  
Fluorescence In Situ Hybridisation ◽  
45S Rdna ◽  
C Banding ◽  
Rdna Sequences ◽  
Rdna Signal ◽  
5S And 45S Rdna

Detailed karyotypes of Lilium longiflorum and L. rubellum were constructed on the basis of chromosome arm lengths, C-banding, AgNO3 staining, and PI-DAPI banding, together with fluorescence in situ hybridisation (FISH) with the 5S and 45S rDNA sequences as probes. The C-banding patterns that were obtained with the standard BSG technique revealed only few minor bands on heterologous positions of the L. longiflorum and L. rubellum chromosomes. FISH of the 5S and 45S rDNA probes on L. longiflorum metaphase complements showed overlapping signals at proximal positions of the short arms of chromosomes 4 and 7, a single 5S rDNA signal on the secondary constriction of chromosome 3, and one 45S rDNA signal adjacent to the 5S rDNA signal on the subdistal part of the long arm of chromosome 3. In L. rubellum, we observed co-localisation of the 5S and 45S rDNA sequences on the short arm of chromosomes 2 and 4 and on the long arms of chromosomes 2 and 3, and two adjacent bands on chromosome 12. Silver staining (Ag-NOR) of the nucleoli and NORs in L. longiflorum and L. rubellum yielded a highly variable number of signals in interphase nuclei and only a few faint silver deposits on the NORs of mitotic metaphase chromosomes. In preparations stained with PI and DAPI, we observed both red- and blue-fluorescing bands at different positions on the L. longiflorum and L. rubellum chromosomes. The red-fluorescing or so-called reverse PI-DAPI bands always coincided with rDNA sites, whereas the blue-fluorescing DAPI bands corresponded to C-bands. Based on these techniques, we could identify most of chromosomes of the L. longiflorum and L. rubellum karyotypes.Key words: fluorescence in situ hybridisation, FISH, 5S rDNA, 45S rDNA, C-banding, reverse PI-DAPI banding.

Chromosomal localization of two novel repetitive sequences isolated from the Chenopodium quinoa Willd. genome

Genome ◽  
2011 ◽  
Vol 54 (9) ◽  
pp. 710-717 ◽  
Author(s):  
B. Kolano ◽  
B.W. Gardunia ◽  
M. Michalska ◽  
A. Bonifacio ◽  
D. Fairbanks ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Repetitive Dna ◽  
Dna Sequences ◽  
Repetitive Sequences ◽  
Chenopodium Quinoa ◽  
5S Rdna ◽  
Rrna Gene ◽  
Fish Analysis ◽  
Rdna Loci

The chromosomal organization of two novel repetitive DNA sequences isolated from the Chenopodium quinoa Willd. genome was analyzed across the genomes of selected Chenopodium species. Fluorescence in situ hybridization (FISH) analysis with the repetitive DNA clone 18–24J in the closely related allotetraploids C. quinoa and Chenopodium berlandieri Moq. (2n = 4x = 36) evidenced hybridization signals that were mainly present on 18 chromosomes; however, in the allohexaploid Chenopodium album L. (2n = 6x = 54), cross-hybridization was observed on all of the chromosomes. In situ hybridization with rRNA gene probes indicated that during the evolution of polyploidy, the chenopods lost some of their rDNA loci. Reprobing with rDNA indicated that in the subgenome labeled with 18–24J, one 35S rRNA locus and at least half of the 5S rDNA loci were present. A second analyzed sequence, 12–13P, localized exclusively in pericentromeric regions of each chromosome of C. quinoa and related species. The intensity of the FISH signals differed considerably among chromosomes. The pattern observed on C. quinoa chromosomes after FISH with 12–13P was very similar to GISH results, suggesting that the 12–13P sequence constitutes a major part of the repetitive DNA of C. quinoa.

Physical mapping of the 5S rDNA gene complex in rice (Oryza sativa)

Genome ◽  
1993 ◽  
Vol 36 (4) ◽  
pp. 658-661 ◽  
Author(s):  
Y. C. Song ◽  
J. P. Gustafson
Keyword(s):  
In Situ Hybridization ◽  
Physical Mapping ◽  
Rice Chromosome ◽  
5S Rdna ◽  
Rice Cultivar ◽  
Chromosome 9 ◽  
Gene Complex ◽  
Rdna Gene ◽  
Biotin Labelling

This study was designed to use biotin labelling in situ hybridization to physically map the 5S rDNA genes to a chromosome arm location in rice. Chromosome preparations were made using an improved protoplast technique, which resulted in more mitotic cells with less overlying cytoplasmic and cellular debris. Cells in which both chromatids were labelled were observed. The hybridization detection level for the 5S rDNA gene complex was 17.22%. The results established that the 5S rDNA gene complex of rice is located at the end of the short arm of chromosome 9 in rice cultivar IR36. The similarities and differences of the 5S rDNA gene complex location between rice and other cereals and advantages of in situ hybridization for physical mapping are discussed.Key words: biotin labelling, in situ hybridization.

Application of fiber-FISH in physical mapping of Arabidopsis thaliana

Genome ◽  
1998 ◽  
Vol 41 (4) ◽  
pp. 566-572 ◽  
Author(s):  
Scott A Jackson ◽  
Ming Li Wang ◽  
Howard M Goodman ◽  
Jiming Jiang
Keyword(s):  
Arabidopsis Thaliana ◽  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Physical Mapping ◽  
Molecular Techniques ◽  
Fish Analysis ◽  
Contig Assembly ◽  
Fish Technique ◽  
Bac Clones

Arabidopsis thaliana has become a model plant species for genetic studies because of its small genome and short juvenility period. However, the small chromosomes of this species are not suitable for classical cytogenetic studies. Here we demonstrate that the fluorescence in situ hybridization (FISH) technique using extended DNA fibers can be a powerful tool in the physical mapping of the A. thaliana genome. Using a refined fiber-FISH technique we were able to measure DNA clusters as long as 1.71 Mb, more than 1% of the A. thaliana genome. Several small DNA loci, including the telomeres and a dispersed repetitive DNA sequence, mi167, were also analyzed with this technique. The results show that without known adjacent DNA markers such small DNA loci cannot be mapped precisely using fiber-FISH. One of the most difficult obstacles in physical mapping by contig assembly is closing the gaps that are present between adjacent contigs. Currently available molecular techniques are not sufficient to accurately estimate the physical sizes of these gaps. We isolated bacterial artificial chromosome (BAC) clones bordering gaps 2 and 3 on the physical contig map of A. thaliana chromosome II. The BAC clones were used in fiber-FISH analysis and the physical sizes of the two gaps were estimated as 31 kb and more than 500 kb, respectively. Thus, we have demonstrated that fiber-FISH is an efficient technique for determining the physical size of gaps on molecular contig maps. Key words: fluorescence in situ hybridization, DNA fibers, physical mapping, genome analysis.

scholarly journals Fluorescence in situ hybridization mapping of translocations and deletions involving the short arm of human chromosome 12 in malignant hematologic diseases

Blood ◽  
1994 ◽  
Vol 84 (10) ◽  
pp. 3473-3482 ◽  
Author(s):  
H Kobayashi ◽  
KT Montgomery ◽  
SK Bohlander ◽  
CN Adra ◽  
BL Lim ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Cell Line ◽  
Yeast Artificial Chromosome ◽  
Artificial Chromosome ◽  
Chromosome 12 ◽  
Fish Analysis ◽  
Translocation Breakpoints ◽  
Hybridization Mapping

Translocations and deletions of the short arm of chromosome 12 [t(12p) and del(12p)] are common recurring abnormalities in a broad spectrum of hematologic malignant diseases. We studied 20 patients and one cell line whose cells contained 12p13 translocations and/or 12p deletions using fluorescence in situ hybridization (FISH) with phage, plasmid, and cosmid probes that we previously mapped and ordered on 12p12–13. FISH analysis showed that the 12p13 translocation breakpoints were clustered between two cosmids, D12S133 and D12S142, in 11 of 12 patients and in one cell line. FISH analysis of 11 patients with deletions demonstrated that the deletions were interstitial rather than terminal and that the distal part of 12p12, including the GDI-D4 gene and D12S54 marker, was deleted in all 11 patients. Moreover, FISH analysis showed that cells from 3 of these patients contained both a del(12p) and a 12p13 translocation and that the affected regions of these rearrangements appeared to overlap. We identified three yeast artificial chromosome (YAC) clones that span all the 12p13 translocation breakpoints mapped between D12S133 and D12S142. They have inserts of human DNA between 1.39 and 1.67 Mb. Because the region between D12S133 and D12S142 also represents the telomeric border of the smallest commonly deleted region of 12p, we also studied patients with a del(12p) using these YACs. The smallest YAC, 964c10, was deleted in 8 of 9 patients studied. In the other patient, the YAC labeled the del(12p) chromosome more weakly than the normal chromosome 12, suggesting that a part of the YAC was deleted. Thus, most 12p13 translocation breakpoints were clustered within the sequences contained in the 1.39 Mb YAC and this YAC appears to include the telomeric border of the smallest commonly deleted region. Whether the same gene is involved in both the translocations and deletions is presently unknown.

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