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Original title:
Single cell expression analysis of genes with potential mrna gradient in mouse oocytes
Authors: Dorosh, Andriy ; Margaryan, Hasmik ; Vodička, Martin ; Ergang, Peter ; Šídová, Monika ; Dvořáková-Hortová, Kateřina
Document type: Papers
Conference/Event: XXIInd Symposium of biology and immunology of reproduction with international participation, Třešť (CZ), 2016-05-26 / 2016-05-28
Year: 2016
Language: eng
Abstract: In frogs, there are clearly visible differently pigmented animal and vegetal poles of the egg determined before fertilization and leading to asymmetrical divisions. Mammalian egg does not show any comparable differentiation and it has been generally accepted that even the individual blastomeres in 2-cell and 4-cell embryos are homogenous. However, recent findings suggest that those blastomeres display different gene expression patterns and might already possess some inclinations to specific cell lineages. We therefore raised a question, whether there could be any mRNA or protein gradients in pre-fertilization oocytes similar to a previously described amphibian egg one. In mammalian eggs, there is a membrane region that is poor in microvilli, cortical granules are absent beneath plasma membrane and sperm cells generally do not bind to this location. This microvilli free region also covers the egg nucleus, and cytoskeleton localization differs markedly to the rest of the cortical space, forming actin –myosin II cortical cap/ring and is considered as animal pole. The purpose of this study was to determine gene products that can be detected at single cell level using qPCR and display gradient like distribution in mature oocytes. We checked expression of 12 selected genes in a pool of 10 oocytes and single mature oocytes. Then, we analysed gene expression in fixed intact oocytes and those undergoing laser capture microdissection procedure (LCMD). Eventually, we have determined six candidate genes for the study of intracellular spatial gene expression in mature mammalian oocytes by subcellular qPCR and in situ hybridization.
Keywords: in situ hybridization; Laser capture microdissection; oocytes; RT-qPCR; single cell expresion analysis
Project no.: ED1.1.00/02.0109, GA 14-05547S
Funding provider: GA ČR
Host item entry: Book of abstracts of XXIInd Symposium of biology and immunology of reproduction
Institution: Institute of Biotechnology AS ČR (web)
Document availability information: Fulltext is available at the institute of the Academy of Sciences.
Original record: http://hdl.handle.net/11104/0262903
Permalink: http://www.nusl.cz/ntk/nusl-260989
The record appears in these collections:
Research > Institutes ASCR > Institute of Biotechnology
Conference materials > Papers
Authors: Dorosh, Andriy ; Margaryan, Hasmik ; Vodička, Martin ; Ergang, Peter ; Šídová, Monika ; Dvořáková-Hortová, Kateřina
Document type: Papers
Conference/Event: XXIInd Symposium of biology and immunology of reproduction with international participation, Třešť (CZ), 2016-05-26 / 2016-05-28
Year: 2016
Language: eng
Abstract: In frogs, there are clearly visible differently pigmented animal and vegetal poles of the egg determined before fertilization and leading to asymmetrical divisions. Mammalian egg does not show any comparable differentiation and it has been generally accepted that even the individual blastomeres in 2-cell and 4-cell embryos are homogenous. However, recent findings suggest that those blastomeres display different gene expression patterns and might already possess some inclinations to specific cell lineages. We therefore raised a question, whether there could be any mRNA or protein gradients in pre-fertilization oocytes similar to a previously described amphibian egg one. In mammalian eggs, there is a membrane region that is poor in microvilli, cortical granules are absent beneath plasma membrane and sperm cells generally do not bind to this location. This microvilli free region also covers the egg nucleus, and cytoskeleton localization differs markedly to the rest of the cortical space, forming actin –myosin II cortical cap/ring and is considered as animal pole. The purpose of this study was to determine gene products that can be detected at single cell level using qPCR and display gradient like distribution in mature oocytes. We checked expression of 12 selected genes in a pool of 10 oocytes and single mature oocytes. Then, we analysed gene expression in fixed intact oocytes and those undergoing laser capture microdissection procedure (LCMD). Eventually, we have determined six candidate genes for the study of intracellular spatial gene expression in mature mammalian oocytes by subcellular qPCR and in situ hybridization.
Keywords: in situ hybridization; Laser capture microdissection; oocytes; RT-qPCR; single cell expresion analysis
Project no.: ED1.1.00/02.0109, GA 14-05547S
Funding provider: GA ČR
Host item entry: Book of abstracts of XXIInd Symposium of biology and immunology of reproduction
Institution: Institute of Biotechnology AS ČR (web)
Document availability information: Fulltext is available at the institute of the Academy of Sciences.
Original record: http://hdl.handle.net/11104/0262903
Permalink: http://www.nusl.cz/ntk/nusl-260989
The record appears in these collections:
Research > Institutes ASCR > Institute of Biotechnology
Conference materials > Papers
Record created 2016-10-15, last modified 2021-11-24