In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models

Abstract Most transgenic animals are generated using a genome-modified stem cell system and genome modification directly in embryos. Although this system is well-established in the development of transgenic animals, donor cell-derived transgenic animal production is inefficient in some cases. Especi...

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Autores principales:	Young Min Kim, Kyung Je Park, Jin Se Park, Kyung Min Jung, Jae Yong Han
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/24fcf1f8897443d8a5fd8682c357cdfb
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spelling	oai:doaj.org-article:24fcf1f8897443d8a5fd8682c357cdfb2021-12-02T16:56:02ZIn vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models10.1038/s41598-021-88706-62045-2322https://doaj.org/article/24fcf1f8897443d8a5fd8682c357cdfb2021-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-88706-6https://doaj.org/toc/2045-2322Abstract Most transgenic animals are generated using a genome-modified stem cell system and genome modification directly in embryos. Although this system is well-established in the development of transgenic animals, donor cell-derived transgenic animal production is inefficient in some cases. Especially in avian models such as chickens, the efficiency of transgenic animal production through primordial germ cells (PGCs) is highly variable compared with embryonic manipulation of mammalian species. Because germ cell and germline-competent stem cell-mediated systems that contain the transgene are enriched only at the upstream level during cell cultivation, the efficiency of transgenic animal production is unreliable. Therefore, we developed an in vivo selection model to enhance the efficiency of transgenic chicken production using microsomal glutathione-S-transferase II (MGSTII)-overexpressing PGCs that are resistant to the alkylating agent busulfan, which induces germ cell-specific cytotoxicity. Under in vitro conditions, MGSTII-tg PGCs were resistant to 1 μM busulfan, which was highly toxic to wild-type PGCs. In germline chimeric roosters, transgene-expressing germ cells were dominantly colonized in the recipient testes after busulfan exposure compared with non-treated germline chimera. In validation of germline transmission, donor PGC-derived progeny production efficiency was 94.68%, and the transgene production rate of heterozygous transgenic chickens was significantly increased in chickens that received 40 mg/kg busulfan (80.33–95.23%) compared with that of non-treated germline chimeras (51.18%). This system is expected to significantly improve the efficiency of generating transgenic chickens and other animal species by increasing the distribution of donor cells in adult testes.Young Min KimKyung Je ParkJin Se ParkKyung Min JungJae Yong HanNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Young Min Kim Kyung Je Park Jin Se Park Kyung Min Jung Jae Yong Han In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
description	Abstract Most transgenic animals are generated using a genome-modified stem cell system and genome modification directly in embryos. Although this system is well-established in the development of transgenic animals, donor cell-derived transgenic animal production is inefficient in some cases. Especially in avian models such as chickens, the efficiency of transgenic animal production through primordial germ cells (PGCs) is highly variable compared with embryonic manipulation of mammalian species. Because germ cell and germline-competent stem cell-mediated systems that contain the transgene are enriched only at the upstream level during cell cultivation, the efficiency of transgenic animal production is unreliable. Therefore, we developed an in vivo selection model to enhance the efficiency of transgenic chicken production using microsomal glutathione-S-transferase II (MGSTII)-overexpressing PGCs that are resistant to the alkylating agent busulfan, which induces germ cell-specific cytotoxicity. Under in vitro conditions, MGSTII-tg PGCs were resistant to 1 μM busulfan, which was highly toxic to wild-type PGCs. In germline chimeric roosters, transgene-expressing germ cells were dominantly colonized in the recipient testes after busulfan exposure compared with non-treated germline chimera. In validation of germline transmission, donor PGC-derived progeny production efficiency was 94.68%, and the transgene production rate of heterozygous transgenic chickens was significantly increased in chickens that received 40 mg/kg busulfan (80.33–95.23%) compared with that of non-treated germline chimeras (51.18%). This system is expected to significantly improve the efficiency of generating transgenic chickens and other animal species by increasing the distribution of donor cells in adult testes.
format	article
author	Young Min Kim Kyung Je Park Jin Se Park Kyung Min Jung Jae Yong Han
author_facet	Young Min Kim Kyung Je Park Jin Se Park Kyung Min Jung Jae Yong Han
author_sort	Young Min Kim
title	In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
title_short	In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
title_full	In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
title_fullStr	In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
title_full_unstemmed	In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
title_sort	in vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/24fcf1f8897443d8a5fd8682c357cdfb
work_keys_str_mv	AT youngminkim invivoenrichmentofbusulfanresistantgermcellsforefficientproductionoftransgenicavianmodels AT kyungjepark invivoenrichmentofbusulfanresistantgermcellsforefficientproductionoftransgenicavianmodels AT jinsepark invivoenrichmentofbusulfanresistantgermcellsforefficientproductionoftransgenicavianmodels AT kyungminjung invivoenrichmentofbusulfanresistantgermcellsforefficientproductionoftransgenicavianmodels AT jaeyonghan invivoenrichmentofbusulfanresistantgermcellsforefficientproductionoftransgenicavianmodels
_version_	1718382786891481088

In vivo enrichment of busulfan-resistant germ cells for efficient production of transgenic avian models

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