Obtainment of chimeric blastocysts of mice by methods of laser nanosurgery

Obtainment of chimeric blastocysts of mice by methods of laser nanosurgery The procedure of obtainment of chimeric blastocysts of mice by laser nanosurgery methods with-out using any other techniques is described. To perform the experiments, a special laser micromanipulator was invented. The murine embryonic stem cells (ESC), which were transformed with pEF-GFP vector, encoding the green fluorescent protein, were used in the experiments. ESC were introduced into the perivitelline space of murine embryos at the stage of 8 cells using the laser micromanipulator. The operated embryos were cultured in vitro until the stage of emergence from zona pellucida. The fluorescence and its precise localization were registered using a confocal microscope. It was shown for the first time that the inclusions of ESC introduced with the lased micromanipulator were found not only in the inner cell mass (ICM) but also in the trophectoderm of the chimeric blastocyst. The technology of nanosurgical operations at early stage preimplanted mammalian embryos using laser techniques opens great opportunities not only for solution of fundamental tasks of experimental embryology of mammals but also for obtainment of chimeric and transgenic animals with predetermined genotype. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Russian Journal of Developmental Biology Springer Journals
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Publisher
Springer US
Copyright
Copyright © 2013 by Pleiades Publishing, Inc.
Subject
Life Sciences; Developmental Biology; Animal Anatomy / Morphology / Histology
ISSN
1062-3604
eISSN
1608-3326
D.O.I.
10.1134/S1062360413060088
Publisher site
See Article on Publisher Site

Abstract

The procedure of obtainment of chimeric blastocysts of mice by laser nanosurgery methods with-out using any other techniques is described. To perform the experiments, a special laser micromanipulator was invented. The murine embryonic stem cells (ESC), which were transformed with pEF-GFP vector, encoding the green fluorescent protein, were used in the experiments. ESC were introduced into the perivitelline space of murine embryos at the stage of 8 cells using the laser micromanipulator. The operated embryos were cultured in vitro until the stage of emergence from zona pellucida. The fluorescence and its precise localization were registered using a confocal microscope. It was shown for the first time that the inclusions of ESC introduced with the lased micromanipulator were found not only in the inner cell mass (ICM) but also in the trophectoderm of the chimeric blastocyst. The technology of nanosurgical operations at early stage preimplanted mammalian embryos using laser techniques opens great opportunities not only for solution of fundamental tasks of experimental embryology of mammals but also for obtainment of chimeric and transgenic animals with predetermined genotype.

Journal

Russian Journal of Developmental BiologySpringer Journals

Published: Nov 27, 2013

References

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