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Production of transgenic rabbit embryos through intracytoplasmic sperm injection

Published online by Cambridge University Press:  27 July 2010

Qiuyan Li ,
Jian Hou ,
Sheng Wang ,
Yongfu Chen  and
Xiao-Rong An
Qiuyan Li
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, Beijing 100094, P.R. China.
Jian Hou
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, Beijing 100094, P.R. China.
Sheng Wang
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, Beijing 100094, P.R. China.
Yongfu Chen
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, Beijing 100094, P.R. China.
Xiao-Rong An*
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, No. 2 Yuan-Ming-Yuan West Road, Haidian District, Beijing 100094, P.R. China.
*
All correspondence to: Xiao-Rong An, State Key Laboratory for Agrobiotechnology, College of Biological Science, China Agricultural University, No. 2 Yuan-Ming-Yuan West Road, Haidian District, Beijing 100094, P.R. China. Tel:/Fax: +86 10 62733463. e-mail: [email protected]
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Summary

The objective of this study was to test if intracytoplasmic sperm injection (ICSI)-mediated gene transfer was an effective method in the production of transgenic rabbit embryos. Rabbit sperm diluted in different media with various pH were treated by freezing without cryoprotectant, and their ability for DNA uptake was determined. In these experiments using production of transgenic rabbit embryos by ICSI, exogenous genes at three concentrations and of two conformation types were used. The rate of DNA association to the sperm seen by rhodamine-tagged DNA encoding green fluorescent protein (GFP) was 90.0%, 92.7%, 91.0%, 91.7%, and 92.3%, respectively in TCM199, DM, DPBS, CZB, and HCZB media. The DNA attachment to sperm was not affected by media pH within the range of 5.4–9.4 (p > 0.05). Expression of GFP first occurred at the 2-cell stage and continued to blastocyst formation. DNA concentration (between 5, 10, and 20 ng/μl) or conformation (linear and circular) had no effect on the production rate of transgenic embryos. These results indicated that genetically modified rabbit blastocysts can be efficiently produced by ICSI technique.

Keywords

Frozen spermICSIRabbitTransgenic embryo
Type
Research Article
Information
Zygote , Volume 18 , Issue 4 , November 2010 , pp. 301 - 307
Copyright
Copyright © Cambridge University Press 2010

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