Identification of chromosomes in Thinopyrum intermedium and wheat Th. intermedium amphiploids based on multiplex oligonucleotide probes

Genome ◽  
2018 ◽  
Vol 61 (7) ◽  
pp. 515-521 ◽  
Author(s):  
Yu Cui ◽  
Yanping Zhang ◽  
Juan Qi ◽  
Honggang Wang ◽  
Richard R.-C. Wang ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fish Analysis ◽  
Oligonucleotide Probes ◽  
Thinopyrum Intermedium ◽  
Chromosome Identification ◽  
Structural Variations ◽  
Molecular Cytogenetic ◽  
Wide Range ◽  
Cytogenetic Technique

Synthesized oligonucleotides (oligos) can be used as effective probes similar to plasmid clones for chromosome identification in fluorescence in situ hybridization (FISH) analysis, making oligo FISH a simpler and more efficient molecular cytogenetic technique for studying plants. In this study, multiplex oligonucleotide probes, including pSc119.2-1, pAs1-4, (GAA)10, (AAC)6, and pTa71, were combined and used in FISH to identify chromosomes in common wheat, Thinopyrum intermedium, and a wheat – Th. intermedium amphiploid TE256-1. In comparison with general FISH probes, signals generated by the multiplex probes were more abundant, colorful, and characteristic. Combining the results of genomic in situ hybridization (GISH) with FISH, Th. intermedium chromosomes and alien chromosomes in TE256-1 could be classified and identified more precisely, especially the J- and Js-genome chromosomes. Moreover, based on the FISH results using multiplex probes, more structural variations in wheat chromosomes of TE256-1 were detected. The results indicated that multiplex oligo probes would have a wide range of application prospects in the creation and identification of wheat – Th. intermedium germplasms.

Differential sensitivity of 16S rRNA targeted oligonucleotide probes used for fluorescence in situ hybridization is a result of ribosomal higher order structure

1996 ◽  
Vol 42 (10) ◽  
pp. 1061-1071 ◽  
Author(s):  
Marc E. Frischer ◽  
Peter J. Floriani ◽  
Sandra A. Nierzwicki-Bauer
Keyword(s):  
In Situ Hybridization ◽  
16S Rrna ◽  
Fluorescence In Situ Hybridization ◽  
Differential Sensitivity ◽  
Probe Design ◽  
Oligonucleotide Probes ◽  
Whole Cells ◽  
Gene Probes ◽  
Wide Range

The use of 16S rRNA targeted gene probes for the direct analysis of microbial communities has revolutionized the field of microbial ecology, yet a comprehensive approach for the design of such probes does not exist. The development of 16S rRNA targeted oligonucleotide probes for use with fluorescence in situ hybridization (FISH) procedures has been especially difficult as a result of the complex nature of the rRNA target molecule. In this study a systematic comparison of 16S rRNA targeted oligonucleotide gene probes was conducted to determine if target location influences the hybridization efficiency of oligonucleotide probes when used with in situ hybridization protocols for the detection of whole microbial cells. Five unique universal 12-mer oligonucleotide sequences, located at different regions of the 16S rRNA molecule, were identified by a computer-aided sequence analysis of over 1000 partial and complete 16S rRNA sequences. The complements of these oligomeric sequences were chemically synthesized for use as probes and end labeled with either [γ-32P] ATP or the fluorescent molecule tetramethylrhodamine-5/-6. Hybridization sensitivity for each of the probes was determined by hybridization to heat-denatured RNA immobilized on blots or to formaldehyde fixed whole cells. All of the probes hybridized with equal efficiency to denatured RNA. However, the probes exhibited a wide range of sensitivity (from none to very strong) when hybridized with whole cells using a previously developed FISH procedure. Differential hybridization efficiencies against whole cells could not be attributed to cell wall type, since the relative probe efficiency was preserved when either Gram-negative or -positive cells were used. These studies represent one of the first attempts to systematically define criteria for 16S rRNA targeted probe design for use against whole cells and establish target site location as a critical parameter in probe design.Key words: 16S rRNA, oligonucleotide probes, in situ hybridization.

scholarly journals Characterization of chromosome constitution in three wheat - Thinopyrum intermedium amphiploids revealed frequent rearrangement of alien and wheat chromosomes

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yu Cui ◽  
Piyi Xing ◽  
Xiaolei Qi ◽  
Yinguang Bao ◽  
Honggang Wang ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Common Wheat ◽  
Wheat Variety ◽  
Wheat Breeding ◽  
Thinopyrum Intermedium ◽  
Structural Variations ◽  
Molecular Marker Analysis ◽  
Partial Amphiploids

Abstract Background Thinopyrum intermedium (2n = 6x = 42) is an important wild perennial Triticeae species exhibiting many potentially favorable traits for wheat improvement. Wheat-Th. intermedium partial amphiploids serve as a bridge to transfer desirable genes from Th. intermedium into common wheat. Results Three octoploid Trititrigia accessions (TE261–1, TE266–1, and TE346–1) with good resistances to stripe rust, powdery mildew and aphids were selected from hybrid progenies between Th. intermedium and the common wheat variety ‘Yannong 15’ (YN15). Genomic in situ hybridization (GISH), fluorescence in situ hybridization (FISH) and multicolor GISH (McGISH) analyses demonstrated that the three octoploid Trititrigia possess 42 wheat chromosomes and 14 Th. intermedium chromosomes. The 14 alien (Th. intermedium) chromosomes belong to a mixed genome consisting of J-, JS- and St-genome chromosomes rather than a single J, JS or St genome. Different types of chromosomal structural variation were also detected in the 1A, 6A, 6B, 2D and 7D chromosomes via FISH, McGISH and molecular marker analysis. The identity of the alien chromosomes and the variationes in the wheat chromosomes in the three Trititrigia octoploids were also different. Conclusions The wheat-Th. intermedium partial amphiploids possess 14 alien chromosomes which belong to a mixed genome consisting of J-, JS- and St- chromosomes, and 42 wheat chromosomes with different structural variations. These accessions could be used as genetic resources in wheat breeding for the transfer of disease and pest resistance genes from Th. intermedium to common wheat.

Fluorescent in Situ Hybridization Analysis at Bone Marrow Biopsy Section in Multiple Myeloma.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4898-4898
Author(s):  
Eun Hae Cho ◽  
Sang-Mi Lee ◽  
Hyeon-Seok Eom ◽  
In-Suk Kim ◽  
Gyeong-Won Lee ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
In Situ Hybridization ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Cytogenetic Study ◽  
Fish Analysis ◽  
Molecular Cytogenetic ◽  
Plasma Cell Neoplasms

Abstract Abstract 4898 Introduction The technique of fluorescence immunophenotyping and interphase cytogenetics as a tool for the investigation of neoplasms has recently been introduced to detect molecular cytogenetic abnormalities in plasma cell myeloma of bone marrow (BM) aspirate. However, in case of sub-optimal BM aspirate or the focal distribution of myeloma in the BM, the plasma cells are significantly lower in the BM aspirate than those of biopsy section. Therefore, we have developed a sensitive fluorescence in situ hybridization (FISH) technique which is combined with immunochemistry and is applicable to BM biopsy section for molecular cytogenetic study of plasma cell neoplasms. Patients and Methods Conventional cytogenetic analysis and FISH results of BM samples of 35 multiple myeloma (MM) patients at the time of diagnosis have been evaluated. The probe for IgH rearrangement has been used for hybridization with myeloma cells coupled with CD138 immunostain at BM biopsy section. Results Nineteen patients (54.3%) had abnormal FISH IgH results in biopsy section, whereas seven (20%) cases had abnormal findings in BM aspirate. FISH IgH analysis at biopsy section revealed various signal patterns and proportions (range 6-87%) of cells with atypical signals out of CD138 positive cells. Among five cases with <10% of plasma cells at BM aspirate, four (80%) had abnormal FISH results at biopsy section, whereas one (20%) had abnormal signals at aspirate. There is no correlation between the proportions of cells with atypical signal corrected by the plasma cell count at BM aspirate and the proportions of cells with atypical signal at biopsy section. Conclusions FISH analysis combined with immunostain which is applied at biopsy section is a highly sensitive and convenient technique to detect and quantify monoclonal plasma cells. It could be used for molecular cytogenetic study in plasma cell neoplasms even though there are less than 10% of plasma cells at BM aspirate and the monitoring of residual disease. Disclosures Kong: National cancer center, Korea: Research Funding.

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.

Fluorescent in situ hybridization (FISH): A useful diagnostic tool for childhood conjunctival melanoma

2021 ◽  
pp. 112067212110307
Author(s):  
Raquel María Moral ◽  
Carlos Monteagudo ◽  
Javier Muriel ◽  
Lucía Moreno ◽  
Ana María Peiró
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Pediatric Population ◽  
Diagnostic Technique ◽  
Histopathological Diagnosis ◽  
Fish Analysis ◽  
Conjunctival Melanoma ◽  
Adequate Treatment ◽  
First Case

Introduction: Conjunctival melanoma is extremely rare in children and has low rates of resolution. Definitive histopathological diagnosis based exclusively on microscopic findings is sometimes difficult. Thus, early diagnosis and adequate treatment are essential to improve clinical outcomes. Clinical case: We present the first case in which the fluorescent in situ hybridization (FISH) diagnostic technique was applied to a 10-year-old boy initially suspected of having amelanotic nevi in his right eye. Based on the 65% of tumor cells with 11q13 (CCND1) copy number gain and 33% with 6p25 (RREB1) gain as measured by the FISH analysis, and on supporting histopathological findings, the diagnosis of conjunctival melanoma could be made. Following a larger re-excision, adjuvant therapy with Mitomycin C (MMC), cryotherapy and an amniotic membrane graft, the patient has remained disease-free during 9 years of long-term follow-up. Case discussion: Every ophthalmologist should remember to consider and not forget the possibility of using FISH analyses during the differential diagnosis of any suspicious conjunctival lesions. Genetic techniques, such as FISH, have led to great advances in the classification of ambiguous lesions. Evidence-based guidelines for diagnosing conjunctival melanoma in the pediatric population are needed to determine the most appropriate strategy for this age group.

Visualization of sporopollenin-containing pathogenic green micro-alga Prototheca wickerhamii by fluorescent in situ hybridization (FISH)

2009 ◽  
Vol 55 (4) ◽  
pp. 465-472 ◽  
Author(s):  
Ryohei Ueno
Keyword(s):  
Cell Wall ◽  
In Situ Hybridization ◽  
Cell Walls ◽  
Fluorescent In Situ Hybridization ◽  
Rapid Identification ◽  
Oligonucleotide Probes ◽  
Logarithmic Growth Phase ◽  
Prototheca Wickerhamii ◽  
Algal Genus

Fluorescent in situ hybridization (FISH) using taxon-specific, rRNA-targeted oligonucleotide probes is one of the most powerful tools for the rapid identification of harmful microorganisms. However, eukaryotic algal cells do not always allow FISH probes to permeate over their cell walls. Members of the pathogenic micro-algal genus Prototheca are characterized by their distinctive cell-wall component, sporopollenin, an extremely tough biopolymer that resists acid and alkaline hydrolysis, enzyme attack, and acetolysis. To our knowledge, there has been no report of the successful permeation by the oligonucleotide probes over the cell walls of unicellular green micro-algae, which contain sporopollenin. The DNA probes passed through the cell wall of Prototheca wickerhamii after treating the algal cells with cetyltrimethylammonium bromide (CTAB). Most cells in the middle logarithmic growth phase culture fluoresced when hybridized with the rRNA-targeted universal probe for eukaryotes, though individual cells included in this culture differed in the level of cell-wall vulnerability to attack by the polysaccharide-degrading enzyme, thus reflecting the different stages of the life cycle. This is the first report regarding the visualization of sporopollenin-containing, green micro-algal cells by FISH.

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