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Biology
小鼠卵巢表观基因组和转录组的细胞特异性配对询问
小鼠卵巢表观基因组和转录组的细胞特异性配对询问
JoVE Journal
Biology
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JoVE Journal Biology
Cell-Specific Paired Interrogation of the Mouse Ovarian Epigenome and Transcriptome

小鼠卵巢表观基因组和转录组的细胞特异性配对询问

Full Text
1,395 Views
12:25 min
February 24, 2023

DOI: 10.3791/64765-v

Sarah R. Ocañas*1,2,3, José V. V. Isola*1,3, Tatiana D. Saccon1, Kevin D. Pham2, Ana J. Chucair-Elliott2, Augusto Schneider4, Willard M. Freeman2,3, Michael B. Stout1,3

1Aging and Metabolism Research Program,Oklahoma Medical Research Foundation, 2Genes & Human Disease Research Program,Oklahoma Medical Research Foundation, 3Oklahoma City Veterans Affairs Medical Center, 4Nutrition College,Federal University of Pelotas

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Please note that some of the translations on this page are AI generated. Click here for the English version.

Overview

This study optimizes the translating ribosome affinity purification (TRAP) and isolation of nuclei tagged in specific cell types (INTACT) methods for paired interrogation of the ovarian transcriptome and epigenome in a mouse model. The approach enables analysis of specific ovarian cell populations without cell sorting, facilitating insights into cell type-specific changes associated with aging.

Key Study Components

Research Area

  • Cell type-specific analysis
  • Ovarian aging
  • Transcriptomic and epigenomic investigation

Background

  • Understanding ovarian aging is crucial for addressing female infertility.
  • Current methods often require cell sorting, which can limit throughput.
  • This technique offers a streamlined way to analyze ovaries from the same mouse.

Methods Used

  • Optimized TRAP and INTACT methods
  • NuTRAP mouse model crossed with Cyp17a1-Cre line
  • Single ovary collection and specific protocols for nuclear isolation

Main Results

  • Enabled paired analysis of transcriptomic and epigenomic data.
  • Uncovered cell type-specific changes in the ovaries related to aging.
  • Validated the potential of this method for broader application across various cell types.

Conclusions

  • The method demonstrates effectiveness in isolating nucleic acids from specific ovarian cell populations.
  • This approach has significant implications for studies focused on female reproductive health.

Frequently Asked Questions

What are the advantages of this method?
It increases throughput, reduces costs, and eliminates the need for specialized cell sorting equipment.
Can this method be applied to other cell types?
Yes, as long as a specific Cre driver is available for the cell type of interest.
What biological effects were observed in the study?
The study identified specific changes in ovarian cells associated with aging, relevant for understanding reproductive senescence.
Why is cell specificity important?
It ensures that the analysis accurately reflects changes within the targeted cell populations, which is crucial for understanding their biological roles.
What preliminary steps are involved in this method?
The method begins with precise collection and homogenization of the ovary, followed by centrifugation to isolate nuclei.
How significant are the findings regarding female infertility?
The findings enhance the understanding of ovarian aging, which is essential for developing treatments for female infertility.

在该协议中,翻译核糖体亲和纯化(TRAP)方法和特定细胞类型(INTACT)方法中标记的细胞核的分离针对使用NuTRAP小鼠模型交叉到Cyp17a1-Cre小鼠系的细胞特异性卵巢转录组和表观基因组的配对询问进行了优化。

该方法提供了一种从同一只小鼠的特定卵巢细胞群中分离核酸的技术,而无需细胞分选。该技术可以帮助确定特定细胞群在各种条件下的变化。例如,随着年龄的增长。

该技术的主要优点是它允许对特定卵巢细胞类型进行配对的表观基因组和转录组学分析,而无需细胞分选。这显著提高了通量,降低了成本,并且在对卵巢组织进行细胞类型特异性分析时无需专用设备。在这里,我们的目标是确定生殖衰老前卵巢衰老发生的细胞类型特异性变化,这对治疗女性不孕症具有意义。

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