Isolation and Identification of Prepubertal Buffalo (Bubalus bubalis) Spermatogonial Stem Cells.

Journal: Asian-Australasian journal of animal sciences
Published Date: Nov 16, 2015

Abstract

Isolation and culture of spermatogonial stem cells (SSCs) are attractive for production of genetic modified offspring. In the present study, buffalo spermatogonial stem-like cells were isolated, cultured and expression pattern of different germ cell marker genes were determined. To recover spermatogonia, testes from age 3 to 7 months of buffalo were decapsulated, and seminiferous tubules were enzymatically dissociated. Two types of cells, immature sertoli cell and type A spermatogonia were observed in buffalo testes in this stage. Germ cell marker genes, OCT3/4 (Pou5f1), THY-1, c-kit, PGP9.5 (UCHL-1) and Dolichos biflorus agglutinin, were determined to be expressed both in mRNA and protein level by reverse transcription polymerase chain reaction and immunostaining in buffalo testes and buffalo spermatogonial stem-like cells, respectively. In the following, when the isolated buffalo buffalo spermatogonial stem-like cells were cultured in the medium supplemented 2.5% fetal bovine serum and 40 ng/mL glial cell-derived neurotrophic factor medium, SSCs proliferation efficiency and colony number were significantly improved than those of other groups (p<0.05). These findings may help in isolation and establishing long term in vitro culture system for buffalo spermatogonial stem-like cells, and accelerating the generation of genetic modified buffaloes.

Authors

  • Wanyou Feng
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Shibei Chen
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Dagiang Do
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Qinyou Liu
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Yanfei Deng
    Animal Reproduction Institute, State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, Guangxi University, Nanning, China.
  • Xiaocan Lei
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Chan Luo
    Animal Reproduction Institute, State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, Guangxi University, Nanning, China.
  • Ben Huang
    State Key Laboratory of Conservation and Untilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530005, China.
  • Deshun Shi
    Animal Reproduction Institute, State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, Guangxi University, Nanning, China. Electronic address: ardsshi@gxu.edu.cn.

Keywords

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