Construction of LAT1 eukaryotic expression vector of C57 mouse and its effect on the Neuro-2a cell
Author:
Affiliation:

Clc Number:

Fund Project:

  • Article
  • |
  • Figures
  • |
  • Metrics
  • |
  • Reference
  • |
  • Related
  • |
  • Cited by
  • |
  • Materials
  • |
  • Comments
    Abstract:

    Objective To construct the lamino acid transporter 1 eukaryotic expression vector of C57 mouse and to express the gene inNeuro-2atumor cells,and explore the effect of LAT1on proliferation and apoptosis of Neuro-2a cell. Methods The full-length LAT1 cDNA was synthesized by RT-PCR and cloned into pcDNA3.1vector to construct recombinant plasmid. The constructed pcDNA3.1-LAT1vector was verified by Enzyme digestion and sequencing and then transfected intomurine Neuro-2acellsby liposome. The transfected cells were selected with G418 and stably expressed strain was constructed. The expression of LAT1 was detected by RT-PCR and western blot. Proliferation was analyzed by MTT, cell cycle and apoptosis were detected by flow cytometric analysis. Results The full-length LAT1 cDNA was amplified successfully and pcDNA3.1-LAT1eukaryoticvector was constructed successfully.Enzyme digestion and sequencing confirmed the sequence was correct.Neuro-2acells were transfected and Stably expressed strain was constructed successfully.MTT showed that the group of transfected restructuring plasmid could significantly affect Neuro-2a cell proliferation more than the control groups (P< 0.05). From the flowcytometric analysis, LAT1 could promote cell proliferation and inhibit Neuro-2a cell apoptosis. Conclusion LAT1 can express successfully in Neuro-2acells which were transfected with recombinant pcDNA3.1-LAT1plasmid.LAT1 in Neuro-2a cells can promote cell proliferation and inhibit the cell apoptosis which provides a basis for the study of LAT1.That lays the foundation for studying biological effects of LAT1.

    Reference
    Related
    Cited by
Get Citation
Share
Article Metrics
  • Abstract:
  • PDF:
  • HTML:
  • Cited by:
History
  • Received:
  • Revised:March 28,2014
  • Adopted:
  • Online: June 11,2014
  • Published: