Related Experiment Video
Updated: Apr 10, 2026
3-D Imaging and Analysis of Neurons Infected In Vivo with Toxoplasma gondii
Published on: December 9, 2014
[Prokaryotic Expression and Immunoreactivity Analysis on Profilin of Toxoplasma gondii]
Objective:
To clone and express the profilin (PRF) gene of Toxoplasma gondii, and analyze the immunoreactivity.
Methods:
Total RNA was extracted from tachyzoites of T. gondii RH strain. The coding region of TgPRF was amplified with a pair of specific primers. PCR product was digested with double restriction enzyme and ligated into pET30a(+) vector. The recombinant pET30a(+)-TgPRF plasmid was transformed into E. coli DH5α with positive clones confirmed by the double restriction enzyme digestion, PCR and sequencing. The correct plasmid was transformed into E. coli BL21 and induced by IPTG. The expressed proteins were purified with Ni-NTA affinity chromatography and analyzed by SDS-PAGE. Western blotting with rabbit anti-T. gondii serum was used to analyze its antigenicity.
Results:
The product of RT-PCR was with 492 bp. pET30a-TgPRF was confirmed by the double restriction enzyme digestion, PCR and sequencing. SDS-PAGE analysis showed that the expressed product was a soluble protein with a relative molecular weight of 35,000. Western blotting assay revealed that rTgPRF was recognized by rabbit anti-T. gondii serum.
Conclusion:
TgPRF gene has been expressed in prokaryotic expression system and shows immunoreactivity.
More Related Videos
11:21Forward Genetics Screens Using Macrophages to Identify Toxoplasma gondii Genes Important for Resistance to IFN-γ-Dependent Cell Autonomous Immunity
Published on: March 12, 2015
09:52Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
Published on: July 12, 2013