Preparation of armored RNA containing M gene of influenza H3N2.
- Author:
Xin-fen YU
1
;
Jing-cao PAN
;
Zhi-cheng HUANG
;
Rong YE
;
Yu KOU
Author Information
- Publication Type:Journal Article
- MeSH: Influenza A Virus, H3N2 Subtype; genetics; Plasmids; RNA, Viral; genetics; Reverse Transcriptase Polymerase Chain Reaction; standards; Viral Matrix Proteins; genetics
- From: Chinese Journal of Experimental and Clinical Virology 2007;21(4):343-345
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo prepare the armored RNA containing M gene of influenza H3N2.
METHODSThe vector pAR-1 was constructed from expression vector pET30b in which the bacteriophage MS2 DNA fragment, containing the genes for maturase and coat protein and the pac site, was inserted. The M gene fragment of influenza A was inserted into the HindIII site downstream of the pac site on the pAR-1, which formed a new recombinant plasmid pAR-2. After the prokaryotic expression was carried out, armored RNA AR-2 containing M gene was obtained. AR-2 was purified, and then was quantified by real time RT-PCR. Moreover, the stability of AR-2 was checked.
RESULTSAR-2 was expressed successfully. AR-2 remained stable under various storage environments. Approximately 8.9 x 10(11) copies of AR-2 particles can be purified from one milliliter of culture.
CONCLUSIONIt showed that AR-2 was stable and RNase-resistant, which, as a virus surrogate, would be used as RT-PCR standards, controls and training or proficiency samples.