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Technological method for promoting ectogenesis of premeiotic female germ cells by using activin A (ActA)

A germ cell and in vitro development technology, applied in the technical field of ActA promoting pre-meiotic female germ cell in vitro development, can solve problems such as little understanding of the regulatory mechanism, and achieve huge social and economic value

Inactive Publication Date: 2013-04-24
QINGDAO AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These problems are difficult to overcome by in vitro culture of germ cells, because so far little is known about the regulatory mechanisms of oocyte growth, formation of gap junctions between follicular granulosa cells and oocytes, and initiation of meiosis in oocytes

Method used

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  • Technological method for promoting ectogenesis of premeiotic female germ cells by using activin A (ActA)
  • Technological method for promoting ectogenesis of premeiotic female germ cells by using activin A (ActA)
  • Technological method for promoting ectogenesis of premeiotic female germ cells by using activin A (ActA)

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Embodiment 1

[0025] (1) Materials and methods

[0026] 1. Isolation of fetal mouse genital ridges

[0027] Fetal mouse ovaries are derived from 12.5dpc pregnant mice, and the judgment of pregnant mice is 0.5dpc. Postnatal mouse ovaries were derived from 12-14 dpp mice. The ovary was isolated from the fetal mouse body by mechanical separation, and transferred to the operating solution of phosphate buffered saline (PBS, pH=7.2)+10% fetal calf serum (FCS), washed 3 times in the operating solution, and used Surgical forceps removed the mesonephros connected with the ovary, in fresh culture medium (Millipore high glucose medium (α-MEM, HyClone company); 5% fetal calf serum (FCS, HyClone company); 1% sodium pyruvate (HyClone company); 1% penicillin streptomycin; 10mIU / ml follicle-stimulating hormone (FSH, sigma company) and the ovary was washed 3 times.

[0028] 2 In vitro culture of fetal mouse genital crest

[0029] The day when the genital ridges were isolated and cultured was set as day ...

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Abstract

The invention relates to a technological method for promoting ectogenesis of premeiotic female germ cells by using activin A (ActA). The method comprises the following steps of: separating 12.5 dpc female fetal rat genital ridges by a mechanical method; cultivating the fetal rat genital ridges in vitro; collecting oocytes; differentiating and developing 12.5 dpc premeiotic germ cells into GV stage oocytes by an in vitro culture method and inducing maturation to obtain mature oocytes; and detecting the quality of the oocytes developed in vitro by an in vitro fertilization (IVF) technology to obtain IVF offspring developed in an early stage. The technological method for promoting the ectogenesis of the premeiotic female germ cells by using the ActA is established and is simple, convenient and practical, so a cellular and molecular mechanism of archecyte ectogenesis and follicle ectogenesis is explored theoretically, high-quality oocytes are provided for patients suffering from ovarian infertility, and huge social value and economic value are created.

Description

Technical field: [0001] The present invention relates to a technical method for promoting the in vitro development of pre-meiotic female germ cells by activin A (ActA), in particular to a method for culturing pre-meiotic germ cells of 12.5 dpc (days after mating) in vitro Methods Differentiate and develop into germinal vesicle (GV) stage oocytes and induce maturation, use in vitro fertilization technology to detect the quality of oocytes formed in vitro, and obtain early-developed in vitro fertilized offspring. It belongs to the field of biomedical technology of reproductive medicine. Background technique: [0002] In previous studies, germ cells derived from pre-meiotic mice failed to resume meiosis and developed to MII stage. In female mouse embryos, 13.5dpc germ cells undergo further DNA replication and enter prophase of the first meiotic division. However, in vitro growth of oocytes derived from fetal germ cells can only enter the diplotene phase of the first meiotic d...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/075
Inventor 张西锋李兰张志鹏沈伟
Owner QINGDAO AGRI UNIV
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