ACTA VETERINARIA ET ZOOTECHNICA SINICA ›› 2014, Vol. 45 ›› Issue (8): 1237-1245.doi: 10.11843/j.issn.0366-6964.2014.08.005

Previous Articles     Next Articles

The Clone of ApoER2 Gene and the Inhibitory Effect of Interference Plasmid ApoER2-siRNA on Selenoprotein Expression in Co-culture Sertoli Cells of Sheep

ZHANG Chun-xiang,ZHAO Hui,ZHANG Guo-lin,GUO Li-na,REN You-she*   

  1. (College of Animal Science and Veterinary Medicine,Shanxi Agricultural University,Taigu 030801,China)
  • Received:2014-02-18 Online:2014-08-23 Published:2014-08-23

Abstract:

The aim of this study was to clone ovine ApoER2 (apolipoprotein E receptor-2),construct the plasmid vector ApoER2-siRNA,and investigate the inhibitory effect of ApoER2 gene silencing on expression of Sel P and PHGPx genes in co-culture Sertoli cells of sheep.The ovine ApoER2 was cloned by RT-PCR.Four pGPU6/GFP/Neo-ApoER2 plasmid expression vectors were constructed,and transfected into the co-culture Sertoli cells of sheep by Lipofectamine 2000TM.The inhibitory effect of ApoER2-shRNA was detected by RT-PCR and Western blotting.The effect of ApoER2-shRNA inhibitory on expression of Sel P and PHGPx genes was detected by RT-PCR.The complete CDS of ApoER2 was 2 490 bp encoding a 892-amino-acid protein.The BLAST analysis showed that the entire nucleotide sequence of ApoER2 gene had 97.7% homology with bovine.ApoER2 mRNA levels in co-culture Sertoli cells were decreased significantly after transfection by the recombinant plasmids located in ApoER2-siRNA-1565 (P<0.01)and ApoER2-siRNA-2567 (P<0.01).Expression of Sel P and PHGPx mRNA were significantly decreased in the ApoER2-siRNA-2567 recombinant plasmid transfection group (P<0.01).The complete CDS of ApoER2 was obtained.Recombinant plasmid pGPU6/GFP/Neo-ApoER2-1565 and pGPU6/GFP/Neo-ApoER2-2567 were successfully constructed with effective inhibitory.Interference of ApoER2 gene decreased the expression of of Sel P and PHGPx mRNA,which could provide the scientific basis for further study of ApoER2 roles in pathway of selenium transport.

CLC Number: