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Isolation and culture of embryonic stem-like cells from pig nuclear transfer blastocysts of different days

Published online by Cambridge University Press:  18 May 2011

Guangyun Tan ,
Linzhu Ren ,
Yongye Huang ,
Xiaochun Tang ,
Yang Zhou ,
Yan Zhou ,
Dong Li ,
Hongxiao Song ,
Hongsheng Ouyang  and
Daxin Pang
Guangyun Tan
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Linzhu Ren
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Yongye Huang
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Xiaochun Tang
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Yang Zhou
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Yan Zhou
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Dong Li
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Hongxiao Song
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Hongsheng Ouyang
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
Daxin Pang*
Affiliation:
Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China.
*
All correspondence to: Daxin Pang. Jilin Province Animal Embryo Engineering Key Laboratory, College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, P.R. China. e-mail: pdx@jlu.edu.cn
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Summary

This study was conducted to establish pig embryonic stem (ES)-like cell lines from nuclear transfer blastocysts. A green fluorescent protein (GFP)-expressing cell line was used as the source of donor cells injected into the enucleated oocytes. Blastocysts were collected at D5 (the fifth day), D7 (the seventh day) and D9 (the ninth day). Differential staining was used to assay the viability and development of blastocysts from the 3 days. The number of inner cell mass (ICM) cells increased from 1.83 ± 0.8 (D5) to 5.37 ± 1.2 (D7) to 7.56 ± 1.5 (D9). The expression profiles of embryonic stem (ES) cell factors (OCT4, SOX2, KLF4 and c-MYC) correlated best with the undifferentiated ES state and were identified by qPCR. The expression of the four factors was increased from D5 to D7, whereas the expression decreased from D7 to D9. We tried to isolate ES-like cells from these embryos. However, ES-like cells from the D7 blastocysts grew slowly and expressed alkaline phosphatase. The cells from the D9 blastocysts grew rapidly but did not express alkaline phosphatase. ES-like cells were not isolated from the D5 blastocysts. These results show that the cells from the D7 embryos are pluripotent but grow slowly. The cells from the D9 embryos grow rapidly but start to lose pluripotency.

Keywords

BlastocystsES cellsNuclear transferPigPluripotent
Type
Research Article
Information
Zygote , Volume 20 , Issue 4 , November 2012 , pp. 347 - 352
Copyright
Copyright © Cambridge University Press 2011

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