Cell-Specific Paired Interrogation of the Mouse Ovarian Epigenome and Transcriptome.
Cell-Specific Paired Interrogation of the Mouse Ovarian Epigenome and Transcriptome.
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DOI：
10.3791/64765
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发表时间：
2023-02-24
期刊：
影响因子：
--
通讯作者：
Stout MB
中科院分区：
文献类型：
--
作者：
Ocañas SR;Isola JVV;Saccon TD;Pham KD;Chucair-Elliott AJ;Schneider A;Freeman WM;Stout MB
评估细胞类型特异性表观基因组学和转录组学变化是理解卵巢衰老的关键。为此，使用新型转基因NuTRAP小鼠模型，对翻译核糖体亲和纯化（TRAP）和特异性细胞类型中标记的细胞核分离（INTACT）方法进行优化，用于随后对细胞特异性卵巢转录组和表观基因组进行配对询问。NuTRAP等位基因的表达受floxed STOP盒的控制，并且可以使用启动子特异性Cre系靶向特定的卵巢细胞类型。由于最近的研究表明卵巢基质细胞参与驱动早衰表型，因此使用Cyp 17 a1-Cre驱动器将NuTRAP表达系统靶向基质细胞。NuTRAP构建体的诱导对卵巢基质成纤维细胞具有特异性，并且从单个卵巢获得用于测序研究的足够的DNA和RNA。NuTRAP模型和方法可用于研究任何卵巢细胞类型与可用的Cre线。在该方案中，使用与Cyp 17 a1-Cre小鼠系杂交的NuTRAP小鼠模型，优化翻译核糖体亲和纯化（TRAP）和分离特定细胞类型中标记的核（INTACT）方法，用于细胞特异性卵巢转录组和表观基因组的配对询问。
Assessing cell-type-specific epigenomic and transcriptomic changes is key to understanding ovarian aging. To this end, optimization of the Translating Ribosome Affinity Purification (TRAP) and Isolation of Nuclei Tagged in Specific Cell Types (INTACT) methods was performed for subsequent paired interrogation of the cell-specific ovarian transcriptome and epigenome, using a novel transgenic NuTRAP mouse model. Expression of the NuTRAP allele is under the control of a floxed STOP cassette and can be targeted to specific ovarian cell types using promoter-specific Cre lines. Since recent studies have implicated ovarian stromal cells in driving premature aging phenotypes, the NuTRAP expression system was targeted to stromal cells using a Cyp17a1-Cre driver. Induction of the NuTRAP construct was specific to ovarian stromal fibroblasts and sufficient DNA and RNA for sequencing studies were obtained from a single ovary. The NuTRAP model and methods presented here can be used to study any ovarian cell type with an available Cre line. In this protocol, the translating ribosome affinity purification (TRAP) and the isolation of nuclei tagged in specific cell types (INTACT) methods were optimized for paired interrogation of the cell-specific ovarian transcriptome and epigenome, using the NuTRAP mouse model crossed to a Cyp17a1-Cre mouse line.
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DOI：
10.1126/science.1247651
发表时间：
2014-02-14
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Jaitin DA;Kenigsberg E;Keren-Shaul H;Elefant N;Paul F;Zaretsky I;Mildner A;Cohen N;Jung S;Tanay A;Amit I
通讯作者：
Amit I
DOI：
10.1038/nrg.2017.15
发表时间：
2017-06
期刊：
Nature reviews. Genetics
影响因子：
--
作者：
Prakadan SM;Shalek AK;Weitz DA
通讯作者：
Weitz DA
DOI：
10.1007/978-1-4939-9240-9_9
发表时间：
2019-01-01
期刊：
SINGLE CELL METHODS: SEQUENCING AND PROTEOMICS
影响因子：
--
作者：
Natarajan, Kedar Nath
通讯作者：
Natarajan, Kedar Nath
影响因子：
3.4
作者：
Ocanas, Sarah R.;Pham, Kevin D.;Freeman, Willard M.
通讯作者：
Freeman, Willard M.
单细胞分辨率的小鼠大脑 DNA 甲基化图谱。
影响因子：
64.8
作者：
Liu H;Zhou J;Tian W;Luo C;Bartlett A;Aldridge A;Lucero J;Osteen JK;Nery JR;Chen H;Rivkin A;Castanon RG;Clock B;Li YE;Hou X;Poirion OB;Preissl S;Pinto-Duarte A;O'Connor C;Boggeman L;Fitzpatrick C;Nunn M;Mukamel EA;Zhang Z;Callaway EM;Ren B;Dixon JR;Behrens MM;Ecker JR
通讯作者：
Ecker JR