Ana Claudia Carstea

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MOUSE INTER-STRAIN MITOCHONDRIA TRANSFER USING PIEZO MICROMANIPULATOR FACILITATED INJECTION

Ana Claudia Carstea, A.T. Bogdan, Elena Daniela Ilie, Roxana Vintila

Abstract
Cytoplasm donation by transferring a small amount (5-15%) of ooplasm to boost mitochondrial number and quality is increasing the energy output necessary to fuel the embryo through pre-implantation development. In this study we have investigated the possibility of transferring fluorescence- labelled mitochondria by ooplasm manipulation using a Piezo micromanipulator facilitated injection. We performed mouse inter-strain mitochondria transfer from C57BL/6 oocytes to CD1 zygots. The mitochondria were fluorescent stained with MitoTracker GreenFM and droplet of approximately 5% of the CD1 oocytes donor ooplasm, were injected into the CD1 zygotes. A control group was injected with only buffer to compare the effect of ooplasma and buffer injection. The embryos mitochondrial heteroplasmy was detected using fluorescence microscopy and we have analyzed the effects of ooplasm injection on "in vitro" development of CD1 zygots. The cleavage, morula and blastocysts developmental rate (30 ± 4.5%, 22 ± 29.7%, 19 ± 25.6%) was significant lower comparing with the control group (53 ± 100%, 45 ± 84.9%, 40 ± 75.4%). The mitochondrial heteroplasmic embryos were associated with subtle, transient effects on preimplantational development. Our results indicate that cytoplasmic mitochondria donation by Piezo micromanipulator injection may be an efficient method for producing heteroplasmic animals, even exerting subtle effects on growth early in life.

Key words: ooplasm, mitochondria, micromanipulator, inter-strain