Preparation and evaluation of MS2 bacteriophage-like particles packaging hepatitis E virus RNA.
Hepatitis E virus (HEV) is the pathogen causing hepatitis E (HE). It arouses global public health concern since it is a zoonotic disease. The objective of this letter is to report a cost-effective internal control prepared for monitoring procedures of HEV reverse transcriptase (RT)-PCR detection. A selected conserved HEV RNA fragment was integrated into the downstream of the truncated MS2 bacteriophage genome based on Armored RNA technology. The resulting MS2-HEV gene harbored by the pET-28b-MS2-HEV plasmid was transformed into E. coli BL21(DE3) for expression analysis by SDS-PAGE. The expression products were purified and concentrated by ultrasonication and ultrafiltration separation. The morphology and stability properties of the virus-like particles (VLPs) were evaluated by electron microscopy scanning and nuclease challenges, respectively. SDS-PAGE results showed that the constructed MS2-HEV gene expressed efficiently and the purity of the VLPs was highly consistent with the result in electron microscopy. Stability evaluation results demonstrated that the prepared VLPs exhibited strong resistance to DNase I and RNase A attacks and also performed long-lasting protection of coated HEV RNA for at least 4 months at -20°C. These data revealed that the prepared VLPs meet the basic requirements of use as internal control material in the HEV RNA amplification assay.
College of Animal Husbandry and Veterinary Medicine, Jinzhou Medical University, No. 5-48 Renmin Street, Jinzhou 121001, China.
Shen
S
Wang
College of Animal Husbandry and Veterinary Medicine, Jinzhou Medical University, No. 5-48 Renmin Street, Jinzhou 121001, China.
Ying
Y
Liu
Animal Quarantine Lab, Inspection and Quarantine Technology Center of Hainan Entry-Exit Inspection and Quarantine Bureau, Haikou 570000, China.
Dandan
D
Li
College of Animal Husbandry and Veterinary Medicine, Jinzhou Medical University, No. 5-48 Renmin Street, Jinzhou 121001, China.
Tiezhong
T
Zhou
College of Animal Husbandry and Veterinary Medicine, Jinzhou Medical University, No. 5-48 Renmin Street, Jinzhou 121001, China gordon402@163.com.
Shenyang
S
Gao
College of Animal Husbandry and Veterinary Medicine, Jinzhou Medical University, No. 5-48 Renmin Street, Jinzhou 121001, China.
Enhui
E
Zha
College of Food Science, Shenyang Agricultural University, No.120 Dongling Road, Shenyang 110866, China.
Xiqing
X
Yue
D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D016751:Hepatitis E / Q000175:diagnosis* / Q000821:virology; D016752:Hepatitis E virus / Q000235:genetics*; D006801:Humans; D017909:Levivirus / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques; D012367:RNA, Viral / Q000235:genetics*; D012194:RNA-Directed DNA Polymerase / Q000032:analysis* / Q000235:genetics*
D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase
Armored RNA technology; MS2 bacteriophage; hepatitis E virus