Distribution of introduced human mitochondrial DNA in early stage mouse embryos

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Abstract

Objective. The aim of study was the analysis of human mitochondrial DNA (mtDNA) distribution among murine blastomeres in the embryos developing after an injection of human mitochondria suspension at the stage of one or two cells is presented.

Material and methods. Mice CBA/C57Black from Rappolovo aged three weeks were used. Zygotes were obtained upon hormonal stimulation of animals and mated with males. 3–10 pL of mitochondrial suspension from HepG2 cells was injected into a zygote or one blastomere of a two-cell embryo. Zygotes or two-cell embryos cultured in M3 medium drops covered with mineral oil in Petri dishes. Upon reaching the two-, four- or eight-cell stage the cultured embryos were separated into blastomeres. The latter were lysed and the total DNA was isolated. Human mtDNA was detected by PCR using species-specific primers.

Results. The development of 2848 mouse embryos was monitored. In 520 embryos that achieved the stage of 2, 4, 8 in proper time the presence of human mtDNA was assayed in each blastomere. Along with murine mtDNA all embryos contained human mitochondrial genome, which is an evidence of artificially modelled heteroplasmy. Not every blastomere of transmitochondrial embryos contained foreign (human) mtDNA. Mathematical elaboration evidenced an uneven distribution of human mtDNA in cytoplasm within the time elapsed between the injection of human mitochondria and the subsequent splitting of the embryo.

Conclusion. The results obtained confirm our previous notion of the presence of 10–11 segregation units of human mtDNA in the total amount of mitochondria (about 5 ∙ 102) injected into an embryo.

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About the authors

Maria E. Kustova

Institute of Experimental Medicine

Email: kusmasha@yandex.ru
ORCID iD: 0000-0002-4149-2895
SPIN-code: 7151-4480

PhD in Biological Sciences, senior researcher of the Department of Molecular Genetics

Russian Federation, Saint Petersburg

Vasilina A. Sokolova

Institute of Experimental Medicine

Email: iva-li@mail.ru
ORCID iD: 0000-0002-9204-2448
SPIN-code: 6348-6616

PhD in Biological Sciences, senior researcher of the Department of Molecular Genetics

Russian Federation, Saint Petersburg

Oksana V. Kidgotko

Institute of Experimental Medicine

Email: oks-kidgotko@yandex.ru
ORCID iD: 0000-0002-2182-7782
SPIN-code: 6348-6616

PhD in Biological Sciences, researcher of the Department of Molecular Genetics

Russian Federation, Saint Petersburg

Mikhail G. Bass

Institute of Experimental Medicine

Email: mgb3@yandex.ru

researcher of the Department of Molecular Genetics

Russian Federation, Saint Petersburg

Faina M. Zakharova

Institute of Experimental Medicine; Saint Petersburg State University

Email: fzakharova@mail.ru
ORCID iD: 0000-0002-9558-3979
SPIN-code: 9699-5744

PhD in Biological Sciences, researcher of the Department of Molecular Genetics; Senior lecturer, Department of Embryologyat

Russian Federation, Saint Petersburg

Vadim B. Vasilyev

Institute of Experimental Medicine; Saint Petersburg State University

Author for correspondence.
Email: vadim@biokemis.ru
ORCID iD: 0000-0002-9707-262X
SPIN-code: 6699-6350

Doctor of Medical Sciences, Head of the Department of Molecular Genetics; Professor of Chair of Fundamental Problems of Medicine and Medical Technology

Russian Federation, Saint Petersburg

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Copyright (c) 2020 Kustova M.E., Sokolova V.A., Kidgotko O.V., Bass M.G., Zakharova F.M., Vasilyev V.B.

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