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Updated: Aug 29, 2026
Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
Published on: July 12, 2013
[Construction and sequencing of recombinant plasmid pcDNA3/GRA1 from Toxoplasma gondii]
Li-ting Cai1, Heng-ping Shu, Li-ping Jiang
1Department of Parasitology, Xiangya School of Medicine, Central South University, Changsha 410078, China.
Objective:
To construct a mammalian expression plasmid pcDNA3/GRA1 to express dense granules antigen-1 (GRA1) of Toxoplasma gondii, and to lay a foundation for further studying the protective immunity of pcDNA3/GRA1 as a DNA vaccine.
Methods:
The GRA1 opening reading frame (ORF) was amplified with two specific primers. The ORF and plasmid pcDNA3 were digested with EcoR I and Xho I respectively and the ORF was ligated into the pcDNA3 at polylinker. The recombinant vector pcDNA3/GRA1 was characterized by PCR, restriction enzyme digestion, and sequencing analysis.
Results:
The expected ORF, 573 bp long, was amplified by PCR, and inserted into plasmid pcDNA3. PCR, restriction enzyme digestion and sequencing analysis showed that pcDNA3/GRA1 contained GRA1 ORF with the right orientation.
Conclusion:
The mammalian expression vector pcDNA3/GRA1 is successfully constructed.
