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A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos

Transgenic mice are important tools for genetic analysis. A current prominent method for producing transgenic mice involves pronuclear microinjection into 1-cell embryos. However, the total transgenic efficiency obtained using this method is less than 10%. Here, we demonstrate that highly efficient...

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Autores principales: SUZUKI, Shinnosuke, TSUKIYAMA, Tomoyuki, KANEKO, Takehito, IMAI, Hiroshi, MINAMI, Naojiro
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Society for Reproduction and Development 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4498375/
https://www.ncbi.nlm.nih.gov/pubmed/25740401
http://dx.doi.org/10.1262/jrd.2014-157
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author SUZUKI, Shinnosuke
TSUKIYAMA, Tomoyuki
KANEKO, Takehito
IMAI, Hiroshi
MINAMI, Naojiro
author_facet SUZUKI, Shinnosuke
TSUKIYAMA, Tomoyuki
KANEKO, Takehito
IMAI, Hiroshi
MINAMI, Naojiro
author_sort SUZUKI, Shinnosuke
collection PubMed
description Transgenic mice are important tools for genetic analysis. A current prominent method for producing transgenic mice involves pronuclear microinjection into 1-cell embryos. However, the total transgenic efficiency obtained using this method is less than 10%. Here, we demonstrate that highly efficient transgenesis in mice can be achieved by cytoplasmic microinjection using a hyperactive piggyBac system. In embryos in which hyPBase mRNA and pPB-CAG-TagRFP DNA were co-injected into the cytoplasm, TagRFP fluorescence was observed after the 2-cell stage; when 30 ng/µl pPB-CAG-TagRFP DNA and 30 ng/µl hyPBase mRNA were co-injected, 94.4% of blastocysts were TagRFP positive. Furthermore, a high concentration of hyPBase mRNA resulted in creation of mosaic embryos in which the TagRFP signals partially disappeared. However, suitable concentrations of injected DNA and hyPBase mRNA produced embryos in which almost all blastomeres were TagRFP positive. Thus, the hyperactive piggyBac transposon system is an easy-to-implement and highly effective method that can contribute to production of transgenic mice.
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spelling pubmed-44983752015-07-13 A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos SUZUKI, Shinnosuke TSUKIYAMA, Tomoyuki KANEKO, Takehito IMAI, Hiroshi MINAMI, Naojiro J Reprod Dev Technology Report Transgenic mice are important tools for genetic analysis. A current prominent method for producing transgenic mice involves pronuclear microinjection into 1-cell embryos. However, the total transgenic efficiency obtained using this method is less than 10%. Here, we demonstrate that highly efficient transgenesis in mice can be achieved by cytoplasmic microinjection using a hyperactive piggyBac system. In embryos in which hyPBase mRNA and pPB-CAG-TagRFP DNA were co-injected into the cytoplasm, TagRFP fluorescence was observed after the 2-cell stage; when 30 ng/µl pPB-CAG-TagRFP DNA and 30 ng/µl hyPBase mRNA were co-injected, 94.4% of blastocysts were TagRFP positive. Furthermore, a high concentration of hyPBase mRNA resulted in creation of mosaic embryos in which the TagRFP signals partially disappeared. However, suitable concentrations of injected DNA and hyPBase mRNA produced embryos in which almost all blastomeres were TagRFP positive. Thus, the hyperactive piggyBac transposon system is an easy-to-implement and highly effective method that can contribute to production of transgenic mice. The Society for Reproduction and Development 2015-02-10 2015-06 /pmc/articles/PMC4498375/ /pubmed/25740401 http://dx.doi.org/10.1262/jrd.2014-157 Text en ©2015 Society for Reproduction and Development http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives (by-nc-nd) License.
spellingShingle Technology Report
SUZUKI, Shinnosuke
TSUKIYAMA, Tomoyuki
KANEKO, Takehito
IMAI, Hiroshi
MINAMI, Naojiro
A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title_full A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title_fullStr A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title_full_unstemmed A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title_short A hyperactive piggyBac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
title_sort hyperactive piggybac transposon system is an easy-to-implement method for introducing foreign genes into mouse preimplantation embryos
topic Technology Report
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4498375/
https://www.ncbi.nlm.nih.gov/pubmed/25740401
http://dx.doi.org/10.1262/jrd.2014-157
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