In vitro synthesis and immunogenicity of Mycrobacterium tuberculosis Ag85B-mRNA vaccine
XIA Min
YANG Xiaolan
YANG Penghui
WANG Xiliang
Abstract：This study was performed to transcript Mycobacterium tuberculosis Ag85 B-mRNA and evaluate its immunogenicity in vitro. The Ag85 B coding sequence was optimized by combining codon preference, GC content and free energy of mRNA secondary structure, while the β globin 3'and 5'UTR sequences were inserted at both ends.The sequence of interest is synthesized and subjected to in vitro transcription, then identified by agarose gel electrophoresis. After cell transfection and expression confirmation in vitro, the Ag85 B-Mrna was used to immunize Balb/c mice with protamine for evaluating specific cellular immune responses. Secondary structure analysis showed that optimized sequence of Ag85 B has higher stability. The stable Ag85 B-Mrna was constructed in vitro. Western blot showed after Ag85 B-Mrna transfection into 293 T cells for 24 and 48 hours, there were clear and specific target protein bands. The results of mouse immunoassay showed that Ag85 B-Mrna could induce high level of IFN-γ against mycobacterial Ag85 B which was responsible for Th1-type immune response. In conclusion, the synthesized stable Ag85 B-Mrna vaccine in vitro has good immunogenicity, which lays a foundation for the improvement of Mrna vaccine research and provides new ideas for the development of tuberculosis vaccine.
Keywords：Ag85BmRNA vaccineSequence optimizationImmune effect
Publication Date：2019-01-01
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：5( 404-408 )