Fluorescence In Situ Hybridization for
            Glycine max
            Metaphase Chromosomes

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.

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Horse Radish Peroxidase Labelled Oligonucleotides and Tyramide Signal Amplification for Fluorescence in Situ Hybridization to Low Abundant Cytokine Mrnas

Microscopy and Microanalysis ◽

10.1017/s143192760000790x ◽

1997 ◽

Vol 3 (S2) ◽

pp. 203-204

Author(s):

Mariette van de Corput ◽

Rob van Gijlswijk ◽

Mark Bobrow ◽

Tom Erickson ◽

Roel Dirks ◽

...

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Signal Amplification ◽

Signal To Noise Ratio ◽

High Specificity ◽

Tyramide Signal Amplification ◽

Horse Radish Peroxidase ◽

Metaphase Chromosomes ◽

Generation Capacity

In recent years, Tyramide Signal Amplification (TSA) has gained acclaim as a very sensitive detection method for immunocytochemsitry and fluorescence in situ hybridization (FISH). To maximally exploit the great signal generation capacity of TSA in mRNA-FISH, minimizing signals emanating from non-specifically bound nucleic acid probe becomes of prime importance, because a specificity check of the signals observed in the cytoplasm is virtually impossible. We reasoned that utilization of synthetic oligonucleotides (ONTs) in stead of commonly used cDNAs or cRNAs would diminish non-specific probe binding and that direct Horse Radish Peroxidase (HRP) labelling of ONTs and TSA would enable their in situ detection.This approach was first tested in metaphase DNA-FISH using chromosome-specific repeats as targets. Using bifunctional crosslinking chemistry and HPLC, 5’-hexylamino oligonucleotides for chromosome specific simple satellite and alphoid sequences were conjugated to HRP and purified. Following 15 - 20 min of situ hybridization of a single HRP-ONT probe to metaphase chromosomes and a direct flurochrome-tyramide detection step, such repeat targets could be visualized with high specificity and excellent signal-to noise ratio.

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Detection of myc translocations in lymphoma cells by fluorescence in situ hybridization with yeast artificial chromosomes

Blood ◽

10.1182/blood.v85.8.2132.bloodjournal8582132 ◽

1995 ◽

Vol 85 (8) ◽

pp. 2132-2138 ◽

Cited By ~ 23

Author(s):

ML Veronese ◽

M Ohta ◽

J Finan ◽

PC Nowell ◽

CM Croce

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Yeast Artificial Chromosome ◽

Human Lymphocytes ◽

Artificial Chromosome ◽

Chromosome 8 ◽

Metaphase Chromosomes ◽

Artificial Chromosomes ◽

Myc Gene

Translocations involving chromosome 8 at band q24 and one of the Ig loci on chromosomes 14q32, 22q11, and 2p11 are the hallmark of Burkitt's lymphoma (BL). It has been previously observed that the exact localization of the breakpoints at chromosome 8q24 can vary significantly from patient to patient, scattering over a distance of more than 300 kb upstream of c-myc and about 300 kb downstream of c-myc. To generate probes for fluorescence in situ hybridization (FISH) that detect most c-myc translocations, we screened a yeast artificial chromosome (YAC) library from normal human lymphocytes by colony hybridization, using three markers surrounding the c-myc gene as probes. We obtained 10 YAC clones ranging in size between 500 and 200 kb. Two nonchimeric clones were used for FISH on several BL cell lines and patient samples with different breakpoints at 8q24. Our results show that the YAC clones detected translocations scattered along approximately 200 kb in both metaphase chromosomes and interphase nuclei. The sensitivity, rapidity, and feasibility in nondividing cells render FISH an important diagnostic tool. Furthermore, the use of large DNA fragments such as YACs greatly simplifies the detection of translocations with widely scattered breakpoints such as these seen in BL.

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Mapping small DNA sequences by fluorescence in situ hybridization directly on banded metaphase chromosomes.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.87.16.6223 ◽

1990 ◽

Vol 87 (16) ◽

pp. 6223-6227 ◽

Cited By ~ 80

Author(s):

Y. S. Fan ◽

L. M. Davis ◽

T. B. Shows

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Dna Sequences ◽

Metaphase Chromosomes

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Use of fluorescence in situ hybridization to study the arrangement of DNA sequences in normal and disease-related chromosomes

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100140117 ◽

1995 ◽

Vol 53 ◽

pp. 748-749

Author(s):

Barbara J. F. Trask ◽

Hillary Massa ◽

Cynthia Friedman ◽

Richard Esposito ◽

Ger van den Engh ◽

...

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Dna Sequences ◽

Single Copy ◽

Two Dimensions ◽

Genetic Maps ◽

Epifluorescence Microscopy ◽

Metaphase Chromosomes ◽

Interphase Nuclei

The sites of specific DNA sequences can be fluorescently tagged by fluorescence in situ hybridization (FISH). Different sequences can be labeled with different fluorochromes so that their arrangement can be studied using epifluorescence microscopy. The distances between points on the same or different chromosomes can be determined easily in a large number of interphase nuclei or metaphase chromosomes. A variety of probe types, ranging from single-copy sequences to highly repeated sequences can be employed. Our work has focussed on the analysis of hybridization patterns in two dimensions using conventional fluorescence microscopy.We have used FISH to study various aspects of genome organization that are difficult to study using other techniques. Examples of these applications will be presented.FISH is now the method of choice for determining the chromosomal location of DNA sequences. DNA sequences can be positioned in the genome with <1:1000 accuracy (to a 3-Mbp region within a 3000-Mbp genome). Through FISH, the cytogenetic, physical and genetic maps of chromosomes can be linked.

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Localization of the (TTAGGG)n telomeric sequence in zebrafish chromosomes

Genome ◽

10.1139/g97-098 ◽

1998 ◽

Vol 41 (1) ◽

pp. 136-138 ◽

Cited By ~ 8

Author(s):

E Gornung ◽

I Gabrielli ◽

L Sola

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Danio Rerio ◽

Hot Spot ◽

Telomeric Sequence ◽

Recombination Site ◽

Metaphase Chromosomes ◽

Sequence Distribution

The distribution of the (TTAGGG)n telomeric sequence in Danio rerio (zebrafish) metaphase chromosomes was studied by fluorescence in situ hybridization. This study continues the analysis of the telomeric sequence distribution in fish and confirms that the (TTAGGG)n sequence is conserved in the telomeres of this group of vertebrates. The hybridization signals were restricted to telomeres in all the individuals analysed. With the degree of resolution currently available, no interstitial sites of the telomeric sequence were detected, which suggests that this type of hot-spot recombination site might be absent in zebrafish chromosomes.

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