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Title 

Mitochondria-targeted DsRed2 protein expression during the early stage of bovine somatic cell nuclear transfer embryo development

 

형질전환 체세포를 이용한 복제수정란에서의 초기 배 발생 시기의 미토콘드리아 DNA 확인

Authors 

H J ParkS H MinH ChoiJ ParkSun-Uk KimS LeeSang-Rae LeeI K KongKyu Tae ChangD B KooD S Lee

Publisher 

Springer Verlag (Germany)

Issue Date 

2016

Citation 

In Vitro Cellular & Developmental Biology-Animal, vol. 52, no. 8, pp. 812-822

Keywords 

Embryonic developmentGene expressionLentiviral infectionMitochondriaSomatic cell nuclear transfer

Abstract 

Somatic cell nuclear transfer (SCNT) has been widely used as an efficient tool in biomedical research for the generation of transgenic animals from somatic cells with genetic modifications. Although remarkable advances in SCNT techniques have been reported in a variety of mammals, the cloning efficiency in domestic animals is still low due to the developmental defects of SCNT embryos. In particular, recent evidence has revealed that mitochondrial dysfunction is detected during the early development of SCNT embryos. However, there have been relatively few or no studies regarding the development of a system for evaluating mitochondrial behavior or dynamics. For the first time, in mitochondria of bovine SCNT embryos, we developed a method for the visualization of mitochondria and expression of fluorescence proteins. To express red fluorescence in mitochondria of cloned embryos, bovine ear skin fibroblasts, nuclear donor, were stably transfected with a vector carrying mitochondria-targeting DsRed2 gene tagged with V5 epitope (mito-DsRed2-V5 tag) using lentivirus-mediated gene transfer because of its ability to integrate in the cell genome and the potential for long-term transgene expression in the transduced cells and their dividing cells. From western blotting analysis of V5 tag protein using mitochondrial fraction and confocal microscopy of red fluorescence using SCNT embryos, we found that the mitochondrial expression of the mito-DsRed2 protein was detected until the blastocyst stage. In addition, according to image analysis, it may be suggested possible use of the system for visualization of mitochondrial localization and evaluation of mitochondrial behaviors or dynamics in early development of bovine SCNT embryos.

ISSN 

1071-2690

Link 

http://dx.doi.org/10.1007/s11626-016-0053-x

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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