Improving the generation of genomic-type transgenic mice by ICSI |
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Authors: | Pedro N Moreira Julio Pozueta Miriam Pérez-Crespo Fernando Valdivieso Alfonso Gutiérrez-Adán Lluís Montoliu |
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Institution: | (1) Department of Molecular and Cellular Biology, Centro Nacional de Biotecnologia (CNB-CSIC), Campus de Cantoblanco, C/Darwin, 3, Madrid, 28049, Spain;(2) Department of Molecular Biology, Centro de Biología Molecular “Severo Ochoa” CBMSO-UAM/CSIC, Campus de Cantoblanco, Madrid, 28049, Spain;(3) Department of Animal Reproduction, INIA, Madrid, 28040, Spain;(4) Present address: Center for Neurobiology and Behavior, Columbia University, New York, NY 10032, USA |
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Abstract: | Transgenes included in genomic-type constructs, such as yeast artificial chromosomes (YAC), P1-derived artificial chromosomes,
or bacterial artificial chromosomes (BAC), are normally correctly expressed, according to the endogenous expression pattern
of the homologous locus, because their large size usually ensures the inclusion of all regulatory elements required for proper
gene expression. The use of these large genomic-type transgenes is therefore the method of choice to overcome most position
effects, commonly associated with standard-type transgenes, and to guarantee the faithful transgene expression. However, in
spite of the different methods available, including pronuclear microinjection and the use of embryonic stem cells as vehicles
for genomic transgenes, the generation of transgenic animals with BACs and, particularly, with YACs can be demanding, because
of the low efficiencies requiring extensive microinjection sessions and/or higher number of oocytes. Recently, we have explored
the use of intracytoplasmic sperm injection (ICSI) into metaphase II oocytes as an alternative method for the generation of
YAC transgenic mice. Our results suggest that the use of transgenic strategies based on ICSI significantly enhances the efficiency
of YAC transgenesis by at least one order of magnitude. |
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Keywords: | Artificial chromosomes Intracytoplasmic sperm injection In vitro fertilisation Sperm-mediated gene transfer Transgene integrity Transgenesis efficiency Transgene transmission |
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