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Mutant clone mapping in normal oesophagus

ABSTRACT: We will use targeted exome sequencing to examine normal appearing epithelium and whole exome and whole genome sequencing of microdissected clones identified by immunostaining Some of the samples will be of low DNA concentration and therefore may require extra rounds of amplification during library prep.

PROVIDER: EGAS00001001874 | EGA |

REPOSITORIES: EGA

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EGAS00001001874-sc-2019-04-03T10:20:36Z - samples

Project description:We will use targeted exome sequencing to examine normal appearing epithelium and whole exome and whole genome sequencing of microdissected clones identified by immunostaining Some of the samples will be of low DNA concentration and therefore may require extra rounds of amplification during library prep. . This dataset contains all the data available for this study on 2019-04-03.

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Mutant clone mapping in oesohagus restricted bait

Project description:We will use targeted exome sequencing to examine normal appearing oesophageal epithelium to determine the mutational burden. DNA will be extracted from epithelia of a defined size and we will overlay positional information from the epithelial pieces to give clone maps over a wide area.

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Mutant clone mapping in normal oesohagus and skin

Project description:We will use targeted exome sequencing to examine normal appearing oesophageal epithelium and skin to determine the mutational burden. DNA will be extracted from epithelia of a defined size and we will overlay positional information from the epithelial pieces to give clone maps over a wide area. This will be done in both oesophageal epithelium and skin

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WGS Mutant clone mapping in normal oesohagus and skin

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Real-time quantitative PCR analysis of human ependymal and ventral horn regions in adult control spinal cord

Project description:Adult human ependymal and ventral horn regions were obtained from postmortem frozen samples by Laser Capture Microdissection. Briefly, Cryostat 25 micron sections from were stained with toluidin blue and both regions microdissected and collected on eppendorf (n=4 for each region). Samples mRNA concentration and purity was assessed by electrophoresis (BioRad Experion HighSensitivity kit, USA). RQI values were lower than 6,5 in every case, so that purification was followed by 2 cycle amplification with a kit designed for highly degraded samples (ExpressArt® TRinucleotide mRNA Amplification Kit; #6299-A15, AmpTec, AMSBIO, UK). After amplification, mRNA concentration and purity was assessed both by electrophoresis (BioRad Experion StSens kit, USA) and by spectrophotometry (Nanodrop, Thermo Scientific, USA). We amplified 3.7-37 ng of total RNA, obtaining between 6 and 21 µg of mRNA after 2 rounds. After collecting samples and studying the RNA integrity and quantity, cDNA of samples was selected for gene expression assays using 384 wells Custom Taqman Low Density Arrays. We built arrays with genes belonging to a profile of stemness or ependymoma (see Garcia-Ovejero et al., 2015, BRAIN).

2015-04-12 | GSE65463 | GEO

Macrodissection versus Microdissection of Rectal Carcinoma: Minor Influence of Stroma to Tumor Gene Profiles

Project description:Background: The molecular determinants of carcinogenesis, tumor progression and patient prognosis can be deduced from simultaneous comparison of thousands of genes by microarray analysis. However, the presence of stroma cells in surgically excised carcinoma tissues might obscure the tumor cell-specific gene expression profiles of these samples. To circumvent this complication, laser microdissection can be performed to separate tumor epithelium from the surrounding stroma and healthy tissue. In this report, we compared RNAs isolated from macrodissected, of which only surrounding healthy tissue had been removed, and microdissected rectal carcinoma samples by microarray analysis in order to determine the most reliable approach to detect the expression of tumor cell-derived genes by microarray analysis. Results: As microdissection yielded low tissue and RNA quantities, extra rounds of mRNA amplification were necessary to obtain sufficient RNA for microarray experiments. These second rounds of amplification influenced the gene expression profiles. Moreover, the presence of stroma cells in macrodissected samples had a minor contribution to the tumor cell gene expression profiles, which can be explained by the observation that more RNA is extracted from tumor epithelial cells than from stroma. Conclusion: These data demonstrate that the more convenient procedure of macrodissection can be adequately used and yields reliable data regarding the identification of tumor cell-specific gene expression profiles. Keywords: macrodissection, microdissection, RNA amplification, stroma, rectal carcinoma

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Project description:BRD4-NUT megadomains is an order of magnitude larger than super-enhancer regions and displays a more continuously enriched profile rather than appearing as a cluster of individual peaks.

2015-07-14 | GSE70869 | GEO

Chip-seq BRD4NUT-BioTAP mapping in 293Trex and 797 cells

2015-07-14 | E-GEOD-70869 | biostudies-arrayexpress

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Project description:Fresh frozen sections of islets obtained by surgery were laser capture microdissected using autofluorescence to guide selection of beta cell areas of the islet. RNA was extracted and amplified with 2 rounds of T7 linear amplification. Two technical replicates were hybridized to Affymetrix U95Av2 arrays.

2014-10-28 | E-MTAB-3042 | biostudies-arrayexpress

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