A novel reagent for removing the zona pellucida of mouse embryos and its preparation method and application

A technology of zona pellucida and mice, applied in the field of embryo development, can solve the problems of low embryo safety and slow processing speed, and achieve the effect of low acidic condition requirements, low cost, and large processing volume

Active Publication Date: 2020-07-28
CHONGQING ACAD OF ANIMAL SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these methods generally have problems such as low safety to embryos and slow processing speed.

Method used

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  • A novel reagent for removing the zona pellucida of mouse embryos and its preparation method and application
  • A novel reagent for removing the zona pellucida of mouse embryos and its preparation method and application
  • A novel reagent for removing the zona pellucida of mouse embryos and its preparation method and application

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Embodiment 1

[0025] A reagent for removing the zona pellucida in mice, pH 5.5, containing baclofen at a concentration of 500 μM. The reagent was prepared by dissolving baclofen in PBS solution, adjusting the pH to 5.5, and the concentration of baclofen was 500 μM.

[0026] Treat mouse embryos with the resulting mouse zona pellucida reagent, comprising the following steps:

[0027] 1. Superovulated mice: Superovulate the mice at 13:00 p.m.: intraperitoneally inject PMSG 10IU / mouse, and 48 hours later, intraperitoneally inject hCG 10IU / mouse. After the hCG injection, the male and female mice were caged together at a ratio of 2:1, and the female mice were checked at 8 o'clock in the morning of the next day, and the fertilized female mice were determined by checking whether there was a vaginal suppository.

[0028] 2. Obtaining embryos: On the morning of the day when the fertilized female mouse was confirmed, the fertilized female mouse was quickly and humanely killed by cervical dislocation....

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Abstract

The invention provides a novel reagent capable of dissolving mouse zona pellucid. The reagent comprises baclofen, wherein the concentration of the baclofen is 300 uM-10mM; and the pH value of the reagent is 5-6. The reagent disclosed by the invention is fast in mouse zona pellucid dissolution, high in pH value and slight in embryo damage and can be applied to in vitro fertilization, in vitro artificial nuclear transfer and other experimental techniques.

Description

technical field [0001] The invention belongs to the field of embryo development, and relates to a reagent for removing the zona pellucida of mouse embryos, a preparation method and application thereof. Background technique [0002] Zona pellucida (Zona pelluccida, ZP) is a layer of cell-free glycoprotein structure surrounding all mammalian oocytes, which plays a key role in egg development, fertilization and pre-implantation embryonic development. The zona pellucida of mice is composed of ZP1, ZP2, and ZP3, which is different from other mammals such as pigs and humans. ZP1, ZP2, and ZP3 are not in a messy shape in the ZP, but have their specific configuration. According to electron microscopy analysis, ZP3 has a repeating grid-like fiber structure, and multiple ZP2·ZP3 heterodimers are connected into a string of bead-like long chains through disulfide bridges; ZP1 connects ZP1 and ZP2 by forming a dimer, The combination of the three forms clathrin with a three-dimensional c...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/073
CPCC12N5/0604C12N2500/33
Inventor 梁浩葛良鹏孙静吴梦
Owner CHONGQING ACAD OF ANIMAL SCI
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