scholarly journals In Situ Sodium Bisulfite Modification of Genomic DNA from Microdissected Melanoma Paraffin-Embedded Archival Tissues

2015 ◽  
Author(s):  
Diego M. Marzese ◽  
Sharon K. Huang ◽  
Dave S. B. Hoon
Keyword(s):  
Genomic Dna ◽  
Sodium Bisulfite ◽  
Bisulfite Modification

C-banding and fluorescence in situ hybridization studies of the wheat–alien hybrid 'Agrotana'

Genome ◽  
1994 ◽  
Vol 37 (3) ◽  
pp. 477-481 ◽  
Author(s):  
Jie Xu ◽  
R. L. Conner ◽  
A. Laroche
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Genomic Dna ◽  
Chromosome Engineering ◽  
C Banding ◽  
Chinese Spring Wheat ◽  
Mother Cells ◽  
Alien Chromatin ◽  
Chromosome Segments

'Agrotana', a wheat-alien hybrid (2n = 56), is a potential source of resistance to common root rot, stem rust, wheat streak mosaic virus, and the wheat curl mite. However, the origin of 'Agrotana', reported to be durum wheat × Agropyron trichophorum (pubescent wheatgrass), is uncertain. The objective of this investigation was to determine the chromosome constitution of 'Agrotana' using C-banding and fluorescence in situ hybridization techniques. The F1 hybrid of 'Agrotana' × 'Chinese Spring' wheat showed 7 I + 21 II in 14.9% of the pollen mother cells, evidence of the presence of the A, B, and D genomes in 'Agrotana'. The hybrid had 16 heavily C-banded chromosomes, namely 4A, and 1-7B of wheat, and a translocation that probably involved wheat chromosomes 2A and 2D. In situ hybridization using biotinylated genomic DNA of Ag. trichophorum cv. Greenleaf blocked with CS DNA failed to identify the alien chromosomes in 'Agrotana', indicating that the alien chromosomes were not likely derived from pubescent wheatgrass. In situ hybridization using labelled wheat genomic DNA blocked with 'Agrotana' DNA revealed that 'Agrotana' had 40 wheat, 14 alien, and 2 (a pair) wheat–alien translocated chromosomes. There was no homology between wheat and the alien chromosomes or chromosome segments involved in the wheat–alien recombinant. Two of the seven pairs of alien chromosomes were homoeologous to each other. The ability to identify alien chromatin in wheat using labelled wheat DNA instead of labelled alien DNA will be particularly useful in chromosome engineering of wheat germplasms having alien chromatin of unknown origin.Key words: wheat–alien hybrid, C-banding, fluorescence in situ hybridization, labelled wheat DNA as probe.

Using genomic slot blot hybridization to assess intergeneric Saccharum x Erianthus hybrids (Andropogoneae – Saccharinae)

Genome ◽  
1997 ◽  
Vol 40 (4) ◽  
pp. 428-432 ◽  
Author(s):  
P. Besse ◽  
C. L. McIntyre ◽  
D. M. Burner ◽  
C. G. de Almeida
Keyword(s):  
Genomic Dna ◽  
Blot Hybridization ◽  
Female Parent ◽  
Saccharum Officinarum ◽  
Rapid Screening ◽  
Hybridization Technique ◽  
Slot Blot ◽  
Saccharum Species ◽  
Slot Blot Hybridization

The use of genomic slot blot hybridization enabled the differentiation of hybrids from selfs in Saccharum × Erianthus intergeneric crosses in which Saccharum was used as the female parent. Based on the genomic in situ hybridization technique, slot blots of DNA from the parents and the progeny were blocked with the Saccharum parent DNA and hybridized with the labelled male Erianthus genomic DNA. This technique allowed a rapid screening for hybrids and was sensitive enough to detect a 1/20 dilution of Erianthus in Saccharum DNA, which should enable the detection of most partial hybrids. The genomic slot blot hybridization technique was shown to be potentially useful for assessing crosses involving Saccharum species with either Old World Erianthus section Ripidium or North American Erianthus (= Saccharum) species. The effectiveness of the technique was assessed on 144 progeny of a Saccharum officinarum × Erianthus arundinaceus cross, revealing that 43% of the progeny were selfs. The importance of this test as a tool to support intergeneric breeding programs is discussed.Key words: slot blot, Erianthus, genomic DNA, Saccharum, sugarcane.

scholarly journals Comprehensive DNA 5-Hydroxymethylation Landscapes in Myelodysplastic Syndromes (MDS)

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2996-2996
Author(s):  
Jia Li ◽  
Yue Wei ◽  
Minjung Lee ◽  
Caleb Class ◽  
Hui Yang ◽  
...  
Keyword(s):  
Genomic Dna ◽  
Research Funding ◽  
Sodium Bisulfite ◽  
Oxidation Products ◽  
Epigenetic Mark ◽  
Epigenetic Alterations ◽  
Bioinformatics Analyses ◽  
Myeloid Malignancies ◽  
Bisulfite Treatment ◽  
Genome Wide

Background: Epigenetic regulators as well as epigenetic marks play an essential role in regulating normal hematopoiesis. Epigenetic alterations are one of the early events reflecting changes in cell identify during the transition of a pre-leukemic condition into a malignant phenotype. Capturing epigenetic alterations at early stages of pre-leukemic conditions could benefit the diagnosis and understanding the disease progression during hematopoietic malignant transformation. DNA 5-hydroxymethylation (5hmC) is the one of the major oxidation products of 5-methycytosine (5mC) mediated by the TET protein family dioxygenase. 5hmC has been reported to be involve in the regulation of chromatin accessibility and gene transcription. The dynamic changes of 5hmC have been reported as a hallmark in myeloid malignancies. In our study, we applied an improved 5hmC profiling method (sCMSIP) developed in our lab to systematically profile 5hmC and evaluate the 5hmC dynamics in patients with myeloid malignancies. Methods: In order to profile 5hmC using low amount of input DNA isolated from patient bone-marrow aspirates, we improved our previously developed anti-CMSIP method to capture 5hmC enriched regions using ultra-low amount of genomic DNA (10-20 ng gDNA). Cytosine-5-methylenesulfonatem (CMS) is the 5hmC derivatives upon sodium bisulfite treatment. A home-made anti-CMS antibody was developed to specifically recognize CMS genome-wide after sodium bisulfite treatment. The traditional anti-CMSIP has been widely used to study the genome-wide 5hmC distribution in various biological systems, including mESC, HSC and immune cells. We further optimized the immunoprecipitation procedure to profile genome-wide 5hmC with ultra-low amount of genomic DNA (sensitive CMSIP, sCMSIP). Using this improved CMS-IP method, we performed genome-wide 5hmC analysis in ~100 individuals including healthy donors (n = 10), patients with MDS (n = 62), AML (n = 4) and CMML (n = 13). In parallel, we performed RRBS (DNA methylation) and RNA-seq (transcriptome) analysis on matched samples. We then performed integrative bioinformatics analyses to unveil the potential diagnostic and prognostic values of 5hmC in myeloid neoplasms. Results and Conclusions: Based on our analysis, we found distinct epigenetic landscapes and transcriptomes between healthy donor and patients with myeloid malignancies. Interestingly, we could further separate patients with MDS into three clusters by comparing their 5hmC landscapes with normal, AML and CMML individuals, suggesting that 5hmC might be a sensitive epigenetic mark to reflect the disease status of MDS. Furthermore, we identified a positive correlation between 5hmC distribution and the blast ratio in paired patients before and after hypomethylating agent (HMA) treatment. These findings suggest that 5hmC might be used as a prognostic marker for HMA treatment. Detailed analyses further elucidate the impaired 5hmC enrichment in various key transcription factors that are essential for HSC function and myeloid lineage specification. Disclosures Garcia-Manero: Amphivena: Consultancy, Research Funding; Helsinn: Research Funding; Novartis: Research Funding; AbbVie: Research Funding; Celgene: Consultancy, Research Funding; Astex: Consultancy, Research Funding; Onconova: Research Funding; H3 Biomedicine: Research Funding; Merck: Research Funding.

A rapid procedure for the isolation of C0t-1 DNA from plants

Genome ◽  
1997 ◽  
Vol 40 (1) ◽  
pp. 138-142 ◽  
Author(s):  
Michael S. Zwick ◽  
Robert E. Hanson ◽  
M. Nurul Islam-Faridi ◽  
David M. Stelly ◽  
Rod A. Wing ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Repetitive Dna ◽  
Copy Number ◽  
Genomic Dna ◽  
Mammalian Species ◽  
Repetitive Dnas ◽  
Rapid Procedure ◽  
Dna Elements ◽  
Repetitive Dna Elements

In situ hybridization (ISH) for the detection of single- or low-copy sequences, particularly large DNA fragments cloned into YAC or BAC vectors, generally requires the suppression or "blocking" of highly-repetitive DNAs. C0t-1 DNA is enriched for repetitive DNA elements, high or moderate in copy number, and can therefore be used more effectively than total genomic DNA to prehybridize and competitively hybridize repetitive elements that would otherwise cause nonspecific hybridization. C0t-1 DNAs from several mammalian species are commercially available, however, none is currently available for plants to the best of our knowledge. We have developed a simple 1-day procedure to generate C0t-1 DNA without the use of specialized equipment.Key words: C0t-1 DNA, in situ hybridization, BACs, plants.

Genomic in situ hybridization to sectioned nuclei shows chromosome domains in grass hybrids

1990 ◽  
Vol 95 (3) ◽  
pp. 335-341
Author(s):  
A.R. Leitch ◽  
W. Mosgoller ◽  
T. Schwarzacher ◽  
M.D. Bennett ◽  
J.S. Heslop-Harrison
Keyword(s):  
In Situ Hybridization ◽  
Genomic Dna ◽  
Microscope Level ◽  
Hordeum Chilense ◽  
Root Tip ◽  
Interphase Nuclei ◽  
Light Microscope Level ◽  
Thin Sections ◽  
Electron Microscopes

In situ hybridization using biotinylated total genomic DNA and avidin detection systems was adapted for examination of thin-sectioned plant material in the light and electron microscopes. Root tip material was preserved prior to sectioning, so that the in vivo disposition of the chromatin was maintained. Use of total genomic DNA from Secale africanum as a probe enabled the chromatin from the two parental genomes in the grass hybrid Hordeum chilense × S. africanum to be distinguished. The biotinylated probe preferentially labelled the chromosomes of S. africanum origin. DNA-DNA hybrids were visualized at the light-microscope level by Texas Red fluorescence and at the electron-microscope level by the enzymic precipitation of DAB (diaminobenzidine) or by colloidal gold particles. The use of thin sections allowed the location of probe hybridization to be established unequivocally in both metaphase and interphase nuclei. Analysis of interphase nuclei showed that chromatin originating from the two parental genomes did not intermix but occupied distinct domains.

The interspecific genome structure of cultivated banana, Musa spp. revealed by genomic DNA in situ hybridization

2000 ◽  
Vol 100 (2) ◽  
pp. 177-183 ◽  
Author(s):  
A. D’Hont ◽  
A. Paget-Goy ◽  
J. Escoute ◽  
F. Carreel
Keyword(s):  
In Situ Hybridization ◽  
Genomic Dna ◽  
Genome Structure ◽  
Musa Spp

scholarly journals Comparison Between Integrated Genomic DNA/RNA Profiling and Fluorescence In Situ Hybridization in the Detection of MYC, BCL-2, and BCL-6 Gene Rearrangements in Large B-Cell Lymphomas

2019 ◽  
Vol 153 (3) ◽  
pp. 353-359 ◽  
Author(s):  
Daniel P Cassidy ◽  
Jennifer R Chapman ◽  
Rafael Lopez ◽  
Kyle White ◽  
Yao-Shan Fan ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
B Cell ◽  
Fluorescence In Situ Hybridization ◽  
Genomic Dna ◽  
Standard Of Care ◽  
Gene Rearrangements ◽  
B Cell Lymphomas ◽  
Rna Profiling ◽  
Large B Cell

Abstract Objectives To compare fluorescence in situ hybridization (FISH) and a commercially available sequencing assay for comprehensive genomic profiling (CGP) to determine the best approach to identify gene rearrangements (GRs) in large B-cell lymphomas (LBCLs). Methods Comparison of standard-of-care FISH assays (including a two-probe approach for MYC; break-apart and fusion probes) and an integrated genomic DNA/RNA sequencing CGP approach on a set of 69 consecutive LBCL cases. Results CGP detected GRs, including those involving MYC (1), BCL-2 (3), and BCL-6 (3), not detected by FISH. FISH detected non–IgH-MYC (4) and BCL-6 (2) GRs that were not detected by CGP. In four instances, standalone CGP or FISH testing would have missed a double-hit lymphoma. Conclusions CGP was superior to FISH in the detection of IgH-MYC rearrangements but was inferior for the detection of non–IgH-MYC rearrangements. Our study demonstrates the rationale for development of a customized approach to identify GRs in LBCLs.

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