Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Induction method of immature oocytes and preparation method of mature oocytes

An oocyte and immature technology, which is used in the induction of immature oocytes and the production of mature oocytes

Pending Publication Date: 2022-05-17
株式会社迪瑟夫
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, in the method described in Non-Patent Document 1, in order to obtain oocytes of primordial follicles from mouse pluripotent stem cells, it takes about 3 weeks to 4 weeks. For animals, in order to produce oocytes from pluripotent stem cells in an in vitro culture system, it is estimated that more than 1 year of culture is required

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Induction method of immature oocytes and preparation method of mature oocytes
  • Induction method of immature oocytes and preparation method of mature oocytes
  • Induction method of immature oocytes and preparation method of mature oocytes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0184] (construction of the carrier)

[0185]From the CAG-DD-hTFAP2C plasmid (Reference 6: "Kobayashi T et al., "Principles of early human development and germ cell program from conserved model systems.", Nature, Vol.546, No.7658, p416-420, 2017 .”) The CAG promoter and destabilization domain (DD) were cloned and inserted into the previously used PiggyBAC vector (reference 7: “Shimamoto S et al., “Hypoxia induces the dormant state in oocytes through expression of Foxo3” , PNAS, https: / / doi.org / 10.1073 / pnas.1817223116, 2019.”), the PB-CAG-DD vector was produced. Next, the cDNAs of eight genes, FIGLA, NOBOX, SOHLH1, LHX8, SUB1, STAT3, TBPL2 and DYNLL1, were amplified from the cDNA of the ovary of a female mouse 13.5 days after fertilization, and the In-Fusion HD cloning kit (Bao Biological company) cloned into the PB-CAG-DD vector. Amplification of each cDNA was carried out by PCR using KOD Fx Neo or KOD Plus Neo DNA polymerase (manufactured by TOYOBO Corporation) according to...

Embodiment 2

[0196] Using mouse iPS cells, induction of differentiation into oocytes was studied in the same manner as ES cells.

[0197] (transfection of vector)

[0198] As iPS cells, mouse BVSC iPS cells prepared using the virus buster established in the paper of Non-Patent Document 1 were used. Using Lipofectamine 2000, the PB-CAG-DD vector containing 8 genes constructed in Example 1 and the hyperactive PBase (hypBase) plasmid were simultaneously transfected into mouse BVSC iPS cells. After culturing at 37°C for 5 days in a serum-free medium supplemented with 2i and LIF to proliferate, a single colony was obtained by selection with puromycin.

[0199] (Induction of differentiation into immature oocytes)

[0200] The iPS cells selected with puromycin were cultured at 37° C. for 5 days in the same manner as in Example 1, thereby inducing differentiation into immature oocytes.

[0201] (production of mature oocytes)

[0202] Next, in the above (transfection of the vector), after the t...

Embodiment 3

[0205] (Identification of Factors Important in Oocyte Formation Genes)

[0206] In order to identify genes that are important factors among the oocyte formation genes to form Figure 4 A total of 26 types of vectors were constructed using the same method as in Example 1 in the manner of combining the genes described in the left figure. Next, each vector was transfected into mouse ES cells (Blimpl-mVenus:Stella-ECFP:Npm2-mCherry(BVSCNmC)) by the same method as in Example 1. After transfection, it was cultured at 37°C for 5 days to proliferate, and ES cells (1×10 5 each) in the above-mentioned oocyte differentiation induction medium with 3×10 4 Ovary somatic cells derived from 12.5-day-old female mice after fertilization were mixed to make agglomerates and cultured at 37°C for 2 days. Next, the obtained aggregate was cultured for 21 days by the same method as in Example 1. For the cultured cells, the number of oocytes formed from each cell line was counted. Furthermore, the...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

A method for inducing an immature oocyte comprises introducing four types of genes comprising FIGLA, NOBOX, LHX8, and TBPL2, or transcripts or expression proteins thereof, into at least one type of cell selected from the group consisting of pluripotent stem cells, ectoderm-like cells, and primordial germ cells. A method for producing a mature oocyte comprises: introducing four types of genes comprising FIGLA, NOBOX, LHX8, and TBPL2, or transcripts or expression proteins thereof, into at least one type of cell selected from the group consisting of pluripotent stem cells, ectoderm-like cells, and primordial germ cells; and co-culturing the introduced cells and the ovarian somatic cells.

Description

technical field [0001] The invention relates to a method for inducing immature oocytes and a method for making mature oocytes. [0002] This application claims priority based on Japanese Patent Application No. 2019-166578 filed in Japan on September 12, 2019, the contents of which are incorporated herein by reference. Background technique [0003] In mammals, totipotency, defined as the ability of one cell to generate individuals, is a peculiar property possessed by single cells. However, it has been difficult to develop such a versatile mechanism even in areas where there is a strong social demand such as infertility treatment. The reason for this is that the oogenesis process that takes place in the fetal ovary cannot be reproduced in vitro. [0004] The inventors previously developed an in vitro culture system for reconstituting oocytes from mouse pluripotent stem cells. Specifically, mouse ES cells or iPS cells were induced to differentiate into primordial germ cell-l...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N5/075C12N5/10C12N15/12
CPCC12N5/0609C12N5/0696C12N5/0611C07K14/47C12N2510/00C12N2502/243C12N2506/02C12N2506/45C12N2501/235C12N2501/60C12N5/10C12N15/85C12N2500/90
Inventor 浜崎伸彦林克彦
Owner 株式会社迪瑟夫
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com