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Related Concept Videos

In-vitro Mutagenesis01:16

In-vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

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Related Experiment Video

Updated: Jun 19, 2026

Use of Freeze-thawed Embryos for High-efficiency Production of Genetically Modified Mice
06:46

Use of Freeze-thawed Embryos for High-efficiency Production of Genetically Modified Mice

Published on: April 2, 2020

An efficient method for generating transgenic mice using NaOH-treated spermatozoa.

Chong Li1, Eiji Mizutani, Tetsuo Ono

  • 1Laboratory for Genomic Reprogramming, Center for Developmental Biology, RIKEN, Kobe, Japan. lichong@cdb.riken.jp

Biology of Reproduction
|October 9, 2009
PubMed
Summary
This summary is machine-generated.

An improved method using NaOH-pretreated sperm for intracytoplasmic sperm injection-mediated transgenesis (ICSI-Tr) efficiently generates transgenic (Tg) mice. This technique enhances gene insertion and offspring rates, simplifying Tg mouse production.

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Generation of Genetically Modified Mice through the Microinjection of Oocytes
10:19

Generation of Genetically Modified Mice through the Microinjection of Oocytes

Published on: June 15, 2017

Area of Science:

  • Genetics and Genomics
  • Reproductive Biology
  • Animal Biotechnology

Background:

  • Transgenic (Tg) animals are crucial for studying exogenous gene functions.
  • Intracytoplasmic sperm injection-mediated transgenesis (ICSI-Tr) is a key method for generating Tg mice.
  • Traditional ICSI-Tr methods using freeze-thawed sperm result in chromosomal damage and low success rates.

Purpose of the Study:

  • To develop an improved and efficient method for generating transgenic mice using ICSI-Tr.
  • To optimize sperm pretreatment for enhanced nuclear integrity and gene insertion efficiency.
  • To increase the success rate of producing transgenic offspring via ICSI-Tr.

Main Methods:

  • Spermatozoa were pretreated with 10 mM NaOH to remove the plasma membrane and tail while preserving nuclear integrity.
  • Pretreated sperm heads were incubated with enhanced green fluorescent protein (EGFP) transgene DNA.
  • Microinjection of treated sperm heads into oocytes followed by embryo transfer to generate Tg mice.

Main Results:

  • Optimal conditions involved incubating sperm with 2 ng/microl EGFP for 10 minutes.
  • 55.6% of injected embryos developed to the blastocyst stage with 56.9% showing EGFP fluorescence.
  • A high rate of 10.2% (12 out of 34) of offspring were confirmed transgenic, suitable for both hybrid and inbred mouse strains.

Conclusions:

  • Simple NaOH pretreatment of spermatozoa significantly improves ICSI-Tr efficiency for transgenic mouse production.
  • This optimized method requires fewer oocytes and offers a more accessible approach to generating Tg mice.
  • The technique facilitates efficient exogenous gene insertion into the host genome, advancing genetic research capabilities.