NGHIÊN CỨU TẠO ARMORED RNA LÀM CHỨNG DƯƠNG CHO REAL-TIME RT-PCR PHÁT HIỆN VIRUS MARBURG

Tóm tắt

Marburg virus is the causative agent of a severe viral hemorrhagic fever in human with high mortality
rate. This study aims to prepare virus-modified particles, based on MS2 bacteriophage, to be used as a
positive control for the diagnosis of Marburg virus by Real-time RT-PCR. To this aim, the target region
of Real-time RT-PCR assay was cloned into the pMC037-HisMS2_PLP_pac plasmid, generating
expression vector pMS2-MARV-NP. MS2-like particles were then produced in E. coli BL21(DE3) and
were purified by Ni-NTA affinity chromatography. Analysis by SDS-PAGE and agarose gel
electrophoresis, transmission electron microscopy, and real-time RT-PCR showed that the His-tagged
Armored RNA particles carrying the target region were successfully assembled and purified at high
concentration (3.9 × 108 copies/µL by RT-qPCR quantification) while DNA plasmid contamination was
negligible. Most importantly, these armored RNA particles were found to be resistant to RNase A.
These findings reveal the potential of Armored RNA technology for rapid preparation of ribonucleaseresistant viral RNA controls.