scholarly journals Physical Map of FISH 5S rDNA and (AG3T3)3 Signals Displays Chimonanthus campanulatus R.H. Chang & C.S. Ding Chromosomes, Reproduces its Metaphase Dynamics and Distinguishes Its Chromosomes

Genes ◽  
2019 ◽  
Vol 10 (11) ◽  
pp. 904
Author(s):  
Xiaomei Luo ◽  
Jingyuan Chen
Keyword(s):  
In Situ Hybridization ◽  
Physical Map ◽  
5S Rdna ◽  
Mitotic Metaphase ◽  
Chromosome Counts ◽  
Metaphase Chromosomes ◽  
High Quality ◽  
High Diversity ◽  
Subtelomeric Regions

Chimonanthus campanulatus R.H. Chang & C.S. Ding is a good horticultural tree because of its beautiful yellow flowers and evergreen leaves. In this study, fluorescence in situ hybridization (FISH) was used to analyse mitotic metaphase chromosomes of Ch. campanulatus with 5S rDNA and (AG3T3)3 oligonucleotides. Twenty-two small chromosomes were observed. Weak 5S rDNA signals were observed only in proximal regions of two chromosomes, which were adjacent to the (AG3T3)3 proximal signals. Weak (AG3T3)3 signals were observed on both chromosome ends, which enabled accurate chromosome counts. A pair of satellite bodies was observed. (AG3T3)3 signals displayed quite high diversity, changing in intensity from weak to very strong as follows: far away from the chromosome ends (satellites), ends, subtelomeric regions, and proximal regions. Ten high-quality spreads revealed metaphase dynamics from the beginning to the end and the transition to anaphase. Chromosomes gradually grew larger and thicker into linked chromatids, which grew more significantly in width than in length. Based on the combination of 5S rDNA and (AG3T3)3 signal patterns, ten chromosomes were exclusively distinguished, and the remaining twelve chromosomes were divided into two distinct groups. Our physical map, which can reproduce dynamic metaphase progression and distinguish chromosomes, will powerfully guide cytogenetic research on Chimonanthus and other trees.

FISH analysis of Zanthoxylum armatum based on oligonucleotides for 5S rDNA and (GAA)6

Genome ◽  
2018 ◽  
Vol 61 (9) ◽  
pp. 699-702 ◽  
Author(s):  
Xiaomei Luo ◽  
Juncheng Liu ◽  
Jingyan Wang ◽  
Wei Gong ◽  
Liang Chen ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
5S Rdna ◽  
Fish Analysis ◽  
Oligonucleotide Probes ◽  
Mitotic Metaphase ◽  
Metaphase Chromosomes ◽  
Zanthoxylum Armatum ◽  
Rdna Loci ◽  
Cytogenetic Studies

Fluorescence in situ hybridization (FISH) using oligonucleotide probes for (GAA)6 (18 bp) and ribosomal DNA (rDNA) (5S rDNA, 41 bp) was applied to analyse Zanthoxylum armatum. (GAA)6 loci were detected on the pericentromeric regions of five chromosome pairs, and 5S rDNA loci were also detected on the pericentromeric regions of another two chromosome pairs. The densities and locations of (GAA)6 and 5S rDNA signals varied between individual chromosomes. High-intensity (GAA)6 signals were detected at the centromeres of two large and two smaller metacentric chromosomes. Relatively strong (GAA)6 signals were detected at the centromeres of two relatively small metacentric chromosomes, although strong 5S rDNA signals were detected at the centromeres of two additional smaller metacentric chromosomes. Weak (GAA)6 signals were detected at the centromeres of four large metacentric chromosomes, whereas weak 5S rDNA signals were detected at the centromeres of two smaller metacentric chromosomes. The remaining chromosomes exhibited no signals. Zanthoxylum armatum had 2n = ∼128. The lengths of the mitotic metaphase chromosomes ranged from 1.22 to 2.34 μm. Our results provide information that may be beneficial for future cytogenetic studies and could contribute to the physical assembly of the Zanthoxylum genome.

Characterization of a tandemly repeated subtelomeric sequence with inverted telomere repeats in maize

Genome ◽  
2009 ◽  
Vol 52 (3) ◽  
pp. 286-293 ◽  
Author(s):  
Jun Li ◽  
Fei Yang ◽  
Jia Zhu ◽  
Shibin He ◽  
Lijia Li
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Tandem Repeats ◽  
Repeated Sequence ◽  
Metaphase Chromosomes ◽  
Single Chromosome ◽  
Amplification Band ◽  
Subtelomeric Sequence ◽  
Subtelomeric Regions

In this study, two complementary telomere primers were applied to a single-primer PCR. A clear amplification band was obtained with one primer, while a smear pattern was seen with the other primer. Sequence analysis of the isolated clones from this specific amplification band revealed that a 412 bp clone designated as MTAS1 shared high homology with a reported subtelomeric sequence (382 bp) from maize ( Zea mays L.), which indicated that this clone was possibly present at subtelomeric regions. The clone MTAS1 displayed a novel structural feature flanked by the forward and inverted telomere repeats. Southern hybridization revealed a ladder of hybridization bands, suggesting that MTAS1 was a tandemly repeated sequence. Fluorescence in situ hybridization results showed that the strong MTAS1 signals were present at the ends of short arms of several long chromosomes, confirming that MTAS1 was a subtelomeric sequence and the high brightness of signals further indicated this cloned sequence was a highly and tandemly repetitive sequence in maize. Fluorescence in situ hybridization with telomeric DNA and MTAS1 as probes on metaphase chromosomes and extended genomic DNA fibers showed that hybridization signals of this clone located adjacent to or overlapped with signals of telomere tandem repeats distributed heterogeneously in subtelomeric regions of several chromosomes and even exhibited differences in two subtelomeres of a single chromosome.

DNA sequence mapping in interphase and metaphase chromosomes by fluorescence in situ hybridization

1992 ◽  
Vol 50 (1) ◽  
pp. 496-497
Author(s):  
Barbara Trask ◽  
Susan Allen ◽  
Anne Bergmann ◽  
Mari Christensen ◽  
Anne Fertitta ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequence ◽  
Dna Sequences ◽  
Dual Band ◽  
Nick Translation ◽  
Metaphase Chromosomes ◽  
Band Pass ◽  
Texas Red ◽  
Fluorescent Spot

Using fluorescence in situ hybridization (FISH), the positions of DNA sequences can be discretely marked with a fluorescent spot. The efficiency of marking DNA sequences of the size cloned in cosmids is 90-95%, and the fluorescent spots produced after FISH are ≈0.3 μm in diameter. Sites of two sequences can be distinguished using two-color FISH. Different reporter molecules, such as biotin or digoxigenin, are incorporated into DNA sequence probes by nick translation. These reporter molecules are labeled after hybridization with different fluorochromes, e.g., FITC and Texas Red. The development of dual band pass filters (Chromatechnology) allows these fluorochromes to be photographed simultaneously without registration shift.

Karyotypes and Distribution of Tandem Repeat Sequences in Brassica nigra Determined by Fluorescence in situ Hybridization

2017 ◽  
Vol 152 (3) ◽  
pp. 158-165 ◽  
Author(s):  
Gui-xiang Wang ◽  
Qun-yan He ◽  
Jiri Macas ◽  
Petr Novák ◽  
Pavel Neumann ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Fluorescence Intensity ◽  
Tandem Repeat ◽  
Brassica Nigra ◽  
Metaphase Chromosomes ◽  
Repeat Sequences ◽  
Tandem Repeat Sequences ◽  
Black Mustard

Whole-genome shotgun reads were analyzed to determine the repeat sequence composition in the genome of black mustard, Brassica nigra (L.) Koch. The analysis showed that satellite DNA sequences are very abundant in the black mustard genome. The distribution pattern of 7 new tandem repeats (BnSAT13, BnSAT28, BnSAT68, BnSAT76, BnSAT114, BnSAT180, and BnSAT200) on black mustard chromosomes was visualized using fluorescence in situ hybridization (FISH). The FISH signals of BnSAT13 and BnSAT76 provided useful cytogenetic markers; their position and fluorescence intensity allowed for unambiguous identification of all 8 somatic metaphase chromosomes. A karyotype showing the location and fluorescence intensity of these tandem repeat sequences together with the position of rDNAs and centromeric retrotransposons of Brassica (CRB) was constructed. The establishment of the FISH-based karyotype in B. nigra provides valuable information that can be used in detailed analyses of B. nigra accessions and derived allopolyploid Brassica species containing the B genome.

Physical mapping of barley genes using an ultrasensitive fluorescence in situ hybridization technique

Genome ◽  
2004 ◽  
Vol 47 (1) ◽  
pp. 179-189 ◽  
Author(s):  
J L Stephens ◽  
S E Brown ◽  
N L.V Lapitan ◽  
D L Knudson
Keyword(s):  
In Situ Hybridization ◽  
Hordeum Vulgare ◽  
Fluorescence In Situ Hybridization ◽  
Linkage Map ◽  
Physical Mapping ◽  
Physical Map ◽  
Primary Objective ◽  
Hybridization Technique ◽  
Hordeum Vulgare L

The primary objective of this study was to elucidate gene organization and to integrate the genetic linkage map for barley (Hordeum vulgare L.) with a physical map using ultrasensitive fluorescence in situ hybridization (FISH) techniques for detecting signals from restriction fragment length polymorphism (RFLP) clones. In the process, a single landmark plasmid, p18S5Shor, was constructed that identified and oriented all seven of the chromosome pairs. Plasmid p18S5Shor was used in all hybridizations. Fourteen cDNA probes selected from the linkage map for barley H. vulgare 'Steptoe' × H. vulgare 'Morex' (Kleinhofs et al. 1993) were mapped using an indirect tyramide signal amplification technique and assigned to a physical location on one or more chromosomes. The haploid barley genome is large and a complete physical map of the genome is not yet available; however, it was possible to integrate the linkage map and the physical locations of these cDNAs. An estimate of the ratio of base pairs to centimorgans was an average of 1.5 Mb/cM in the distal portions of the chromosome arms and 89 Mb/cM near the centromere. Furthermore, while it appears that the current linkage maps are well covered with markers along the length of each arm, the physical map showed that there are large areas of the genome that have yet to be mapped.Key words: Hordeum vulgare, barley, physical mapping, FISH, cDNA, genetics, linkage, chromosome, BACs.

Optimization of in situ hybridization to human metaphase chromosomes

1989 ◽  
Vol 182 (1) ◽  
pp. 25-31 ◽  
Author(s):  
Bodil Lomholt ◽  
Pernille Dissing Sørensen ◽  
Henrik Simonsen ◽  
Sune Frederiksen
Keyword(s):  
In Situ Hybridization ◽  
Metaphase Chromosomes

The use of double fluorescence in situ hybridization to physically map the positions of 5S rDNA genes in relation to the chromosomal location of 18S–5.8S–26S rDNA and a C genome specific DNA sequence in the genus Avena

Genome ◽  
1996 ◽  
Vol 39 (3) ◽  
pp. 535-542 ◽  
Author(s):  
Concha Linares ◽  
Juan González ◽  
Esther Ferrer ◽  
Araceli Fominaya
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequence ◽  
Diploid Species ◽  
5S Rdna ◽  
Rdna Genes ◽  
26S Rdna ◽  
A Genome ◽  
Rdna Loci ◽  
C Genome

A physical map of the locations of the 5S rDNA genes and their relative positions with respect to 18S–5.8S–26S rDNA genes and a C genome specific repetitive DNA sequence was produced for the chromosomes of diploid, tetraploid, and hexaploid oat species using in situ hybridization. The A genome diploid species showed two pairs of rDNA loci and two pairs of 5S loci located on both arms of one pair of satellited chromosomes. The C genome diploid species showed two major pairs and one minor pair of rDNA loci. One pair of subtelocentric chromosomes carried rDNA and 5S loci physically separated on the long arm. The tetraploid species (AACC genomes) arising from these diploid ancestors showed two pairs of rDNA loci and three pairs of 5S loci. Two pairs of rDNA loci and 2 pairs of 5S loci were arranged as in the A genome diploid species. The third pair of 5S loci was located on one pair of A–C translocated chromosomes using simultaneous in situ hybridization with 5S rDNA genes and a C genome specific repetitive DNA sequence. The hexaploid species (AACCDD genomes) showed three pairs of rDNA loci and six pairs of 5S loci. One pair of 5S loci was located on each of two pairs of C–A/D translocated chromosomes. Comparative studies of the physical arrangement of rDNA and 5S loci in polyploid oats and the putative A and C genome progenitor species suggests that A genome diploid species could be the donor of both A and D genomes of polyploid oats. Key words : oats, 5S rDNA genes, 18S–5.8S–26S rDNA genes, C genome specific repetitive DNA sequence, in situ hybridization, genome evolution.

Single-copy and amplified CAD genes in Syrian hamster chromosomes localized by a highly sensitive method for in situ hybridization

1982 ◽  
Vol 2 (3) ◽  
pp. 308-319
Author(s):  
G M Wahl ◽  
L Vitto ◽  
R A Padgett ◽  
G R Stark
Keyword(s):  
In Situ Hybridization ◽  
Syrian Hamster ◽  
Specific Radioactivity ◽  
Large Chromosome ◽  
Hybridization Technique ◽  
Wild Type ◽  
Aspartate Transcarbamylase ◽  
Metaphase Chromosomes ◽  
Cad Gene

Syrian hamster cells resistant to N-(phosphonacetyl)-L-aspartate (PALA), a specific inhibitor of the aspartate transcarbamylase activity of the multifunctional protein CAD, overproduce this protein as a result of amplification of the CAD gene. We have used a sensitive in situ hybridization technique to localize CAD genomes in spreads of metaphase chromosomes from several independent PALA-resistant lines and from wild-type PALA-sensitive cells. The amplified genes were always found within chromosomes, usually in an expanded region of the short arm of chromosome B9. In wild-type cells, the CAD gene was also on the short arm of chromosome B9. In one mutant line, 90 to 100 CAD genes were found within an expanded B9 chromosome and 10 to 15 more were near the distal end of one arm of several different chromosomes. Another line contained most the genes in a telomeric chromosome or large chromosome fragment. The amplified genes were in chromosomal regions that were stained in a banded pattern by trypsin-Giemsa. A few double minute chromosomes were observed in a very small fraction of the total spreads examined. The it situ hybridizations were performed in the presence of 10% dextral sulfate 500, which increases the signal by as much as 100-fold. Using recombinant DNA plasmids nick-translated with [125I]dCTP to high specific radioactivity, 10 CAD genes in a single chromosomal region were revealed after 1 week of autoradiographic exposure, and the position of the unique gene could be seen after 1 month.

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