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Quality of transgenic rabbit embryos with different EGFP gene constructs

Published online by Cambridge University Press:  28 May 2010

P. Chrenek ,
M. Bauer  and
A.V. Makarevich
P. Chrenek*
Affiliation:
Animal Production Research Centre Nitra, Hlohovecka 2, 951 41 Luzianky; or Slovak University of Agriculture, Nitra, Slovak Republic.
M. Bauer
Affiliation:
Animal Production Research Centre Nitra, Hlohovecka 2, 951 41 Luzianky, Slovak Republic. Department of Botany and Genetics, Constantine the Philosopher University, Nitra, Slovak Republic.
A.V. Makarevich
Affiliation:
Animal Production Research Centre Nitra, Hlohovecka 2, 951 41 Luzianky, Slovak Republic.
*
All correspondence to Peter Chrenek. Animal Production Research Centre Nitra, Hlohovecka 2, 951 41 Luzianky; or Slovak University of Agriculture, Nitra, Slovak Republic. Tel: +421 37 654 6285. Fax: +421 37 654 6189. e-mail: chrenekp@yahoo.com
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Summary

The aim of this study was to compare the quality of rabbit transgenic embryos obtained upon microinjection of gene constructs containing different promoters and green fluorescent proteins (CMVIE–EGFP, PGK–EGFP and CMVIE–hrGFP). Developmental rate, total cell number in hatching blastocyst stage, number of apoptotic cells, diameter of embryos, transgene integration and transgenic mosaicism were investigated.

The rate of rabbit embryos microinjected with the different gene constructs developed up to morula stage was significantly lower (p < 0.05) than that of intact (non-microinjected) rabbit embryos (66–74vs. 98%). The highest efficiency of transgene integration (15%) was found when the CMVIE–EGFP (DrdI) gene construct was used, however a significantly higher transgenic mosaicism (60%) was found in rabbit embryos using this gene. The lowest cell number was counted in rabbit transgenic embryos with CMVIE–rhGFP linearized by ScaI (115.0 ± 8.20), the highest cell number (134.0 ± 35.00) was detected in rabbit transgenic embryos carrying PGK–EGFP (Not I) gene. The highest number of apoptotic cells (2.6 ± 0.33) was recorded in rabbit transgenic embryos with the integrated CMVIE–EGFP (ClaI) transgene.

Based on these results a more suitable gene marker for rabbit transgenic embryos production and selection is the CMVIE–EGFP (ClaI) gene construct. Prior to using microinjected embryos (for embryo transfer, vitrification or ESC isolation) it is necessary to pre-select microinjected embryos with evident transgenic mosaicism.

Keywords

EmbryoGFPMosaicismQualityRabbitTransgenic
Type
Research Article
Information
Zygote , Volume 19 , Issue 1 , February 2011 , pp. 85 - 90
Copyright
Copyright © Cambridge University Press 2010

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