Prolonged Expression of Exogenous GFP Gene in the Porcine Embryos generated by Intracytoplasmic Sperm Injection-Mediated Gene Transfer

서지정보

ㆍ발행기관 : 한국수정란이식학회 ㆍ수록지정보 : Journal of Embryo Transfer / 30권 / 3호
ㆍ저자명 : Hak-Jae Chung, NaRae Son, Joo-Hee Han, Chun-Gyu Park, Kyung-Woon Kim, Mi-Ryung Park, In-Sul Hwang, Jin-Ki Park, Gi-Sun Im

목차

INTRODUCTION
MATERIALS AND METHODS
1. In Vitro Maturation of Oocytes
2. Sperm Collection and Treatments
3. Intracytoplasmic Sperm Injection and Oocyte Activation
4. Apoptosis Analysis and Cell Counting
5. Embryo Transfer and Recovery
6. Statistics
RESULTS AND DISCUSSION
REFERENCES

영어 초록

Understanding the behavior of transgenes introduced into oocyte or embryos is essential for evaluating the methodologies for transgenic animal production. To date, many studies have reported the production of transgenic pig embryos with, however, low efficiency in environment of blastocyst production. The aim of present study was to determine the expression and duration of transgene transferred by intracytoplasmic sperm injection-mediated gene transfer (ICSI-MGT). Embryos obtained from the ICSI-MGT procedure were analysed for the expression of GFP and then for the transmission of the transgene. Briefly, fresh spermatozoa were bound to exogenous DNA after treatment by Triton X-100 and Lipofectin. When ICSI-MGT was performed using sperm heads with tails removed, the yield of blastocyst (25.3%), treated with Lipofectin (18.8%) and Triton X-100 (19.2%) were observed. Treatments of Lipofectin or Triton X-100 did not further improve the rates of blastocysts. Moreover, the apoptosis rates of embryos were obtained from the control and LIpofectin groups (8.7%, 9.7%, respectively), but were significantly higher in the Triton X-100 group (13.0%). Our results demonstrated that ICSI-MGT caused minimal damage to oocytes that could develop to full term. Moreover, the embryos derived by ICSI-MGT have shown prolonged exogenous DNA expression during preimplantation stage in vivo. However, more efforts will be required to improve the procedures of both sperm treatments cause of high frequency of mosaicisms.

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