Chromosomal Variation in Crocus vernus Hill (Iridaceae) Investigated by in situ Hybridization of rDNA and a Tandemly Repeated Sequence

A tandemly repeated sequence isolated from a clone (HAG004N15) of a nebulized genomic DNA library of sunflower (Helianthus annuus L., 2n = 34) was characterized and used to study the chromosome complement of sunflower. HAG004N15 repeat units (368 bp in length) were found to be highly methylated, and their copy number per haploid (1C) genome was estimated to be 7800. After in situ hybridization of HAG004N15 repeats onto chromosome spreads, signals were observed at the end of both chromosome arms in 4 pairs and at the end of only one arm in 8 other pairs. Signals were also observed at the intercalary (mostly subtelomeric) regions in all pairs, in both arms in 8 pairs, and in only one arm in the other 9 pairs. The short arm of 1 pair was labelled entirely. The chromosomal location of ribosomal DNA was also studied by hybridizing the wheat ribosomal probe pTa71. Four chromosome pairs contained ribosomal cistrons at the end of their shorter arm, but a satellite was seen in only 3 pairs. These hybridization patterns were the same in the 3 sunflower lines studied (HA89, RA20031, and HOR). The chromosomal localization of HAG004N15-related sequences allowed all of the chromosome pairs to be distinguished from each other, in spite of small size and similar morphology.

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Identification of the E3900 family, a second family of rye B chromosome specific repeated sequences

Genome ◽

10.1139/g93-095 ◽

1993 ◽

Vol 36 (4) ◽

pp. 706-711 ◽

Cited By ~ 41

Author(s):

Richard Blunden ◽

Timothy J. Wilkes ◽

John W. Forster ◽

Mar M. Jimenez ◽

Michael J. Sandery ◽

...

Keyword(s):

In Situ Hybridization ◽

Secale Cereale ◽

Repeated Sequence ◽

B Chromosome ◽

Repeated Sequences ◽

Genomic Hybridization ◽

The Family

A second family of highly repeated sequences has been identified on the B chromosome of rye (Secale cereale). The E3900 family was detected as a variant band in EcoRI digests of +B DNA. A clone of the basic repeat of the family was obtained, and the organization of the family was investigated by genomic hybridization. The E3900 family has no apparent homology to the A chromosome sequences of rye or other members of the Gramineae. The family has been localized by in situ hybridization to the end of the long arm of the rye B chromosome. The previously characterized E1100 sequence shows in situ hybridization to the same location as the E3900 family. These results are discussed in light of current theories of the origin of B chromosomes.Key words: B chromosome, Secale cereale, repeated sequence, cloning, in situ hybridization.

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Molecular evidence on the origin of wheat chromosomes 4A and 4B

Genome ◽

10.1139/g90-006 ◽

1990 ◽

Vol 33 (1) ◽

pp. 30-39 ◽

Cited By ~ 16

Author(s):

J. Dvořák ◽

P. Resta ◽

R. S. Kota

Keyword(s):

In Situ Hybridization ◽

Repeated Sequence ◽

Triticum Aestivum L ◽

Close Relative ◽

Molecular Evidence ◽

Substitution Lines ◽

Genome Chromosome ◽

B Genome ◽

A Genome

The genome allocation of the Triticum aestivum L. chromosomes denoted 4A and 4B was based on an erroneous inference. Since neither chromosome pairs with the chromosomes of putative ancestors of wheat, molecular tools were employed to clarify the origin of the two chromosomes. Disomic substitutions for T. aestivum chromosomes 4A or 4B by chromosomes 4 from T. speltoides (Tausch) Gren., a putative ancestor of the wheat B genome, T. longissimum (Schweinf. et Muschl.) Bowden (a close relative of T. speltoides), or T. monococcum L. ssp. aegilopoides (Link) Thell., a close relative of the ancestor of the wheat A genome, were produced. The ability of the substituted chromosome to compensate in the disomic substitution lines, the C-banding patterns of the chromosomes, electrophoretic alleles at the Adh-1 and Lpx-1 loci, and in situ hybridization with an interspersed repeated sequence all were consistent in showing that the chromosome previously denoted as 4A belongs to the B genome and the chromosome previously denoted as 4B is a rearranged chromosome of the A genome. Chromosome 4A is consequently reallocated to the B genome and chromosome 4B to the A genome in T. turgidum L. em. Morris et Sears and T. aestivum. To reflect the fact that the chromosome previously denoted as 4B has only a homoeologous relationship to chromosome 4A of T. urartu (the ancestor of the A genome in polyploid wheats), the chromosome is designated 4Aa.Key words: repeated nucleotide sequence, alcohol dehydrogenase, lipoxygenase, in situ hybridization, chromosome evolution.

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Variation in telomeric heterochromatin in somaclones of rye

Genome ◽

10.1139/g92-088 ◽

1992 ◽

Vol 35 (4) ◽

pp. 590-593 ◽

Cited By ~ 12

Author(s):

A. Karp ◽

P. G. Owen ◽

S. H. Steele ◽

P. J. Bebeli ◽

P. J. Kaltsikes

Keyword(s):

In Situ Hybridization ◽

Somaclonal Variation ◽

Dna Sequences ◽

Repeated Sequence ◽

Interstitial Site ◽

Nucleolus Organizer ◽

Selfed Progeny ◽

Submetacentric Chromosome ◽

Telomeric Heterochromatin

Somaclonal variation in telomeric heterochromatin was detected by in situ hybridization with the het 1 probe, which hybridizes to a 380-bp repeated sequence family. In a control cultivar, Gazelle, large blocks of signal were detected at the telomeres but not at the centromeres or the secondary constrictions. In the donor line, 7R–, labelling was restricted to small telomeric dots, confirming that the large telomeric blocks had been removed in selection of this line. In situ hybridization with the het 1 probe to chromosomes of selfed progeny from 50 plants regenerated from independent cultured immature embryos of the 7R– line revealed variant patterns for three regenerants. In the progeny of two regenerants, a new interstitial hybridization site was detected on the short arm of a submetacentric chromosome. This site was not at the nucleolus organizer. In the progeny of the third regenerant two changes were detected: an enlarged telomeric block on the long arm of an unidentified chromosome and an interstitial site on the long arm of chromosome 6. All three regenerated plants had shown normal morphology and meiotic behaviour. The identification of somaclonal variants in telomeric heterochromatin provides further evidence for variation in repeated DNA sequences in plant tissue culture.Key words: rye (Secale cereale), somaclonal variation, in situ hybridization, repeated sequence, heterochromatin, telomere.

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

Genome ◽

10.1139/g09-005 ◽

2009 ◽

Vol 52 (3) ◽

pp. 286-293 ◽

Cited By ~ 11

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.

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

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100122885 ◽

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.

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Ultrastructural analysis of the spatial distribution of MRNA

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100123167 ◽

1992 ◽

Vol 50 (1) ◽

pp. 552-553

Author(s):

Gary Bassell ◽

Robert H. Singer

Keyword(s):

Electron Microscopy ◽

Spatial Distribution ◽

In Situ Hybridization ◽

High Resolution ◽

Nucleic Acids ◽

Colloidal Gold ◽

Dna Probe ◽

Nucleotide Analogs ◽

Gold Particles

We have been investigating the spatial distribution of nucleic acids intracellularly using in situ hybridization. The use of non-isotopic nucleotide analogs incorporated into the DNA probe allows the detection of the probe at its site of hybridization within the cell. This approach therefore is compatible with the high resolution available by electron microscopy. Biotinated or digoxigenated probe can be detected by antibodies conjugated to colloidal gold. Because mRNA serves as a template for the probe fragments, the colloidal gold particles are detected as arrays which allow it to be unequivocally distinguished from background.

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