[Preparation of armored RNA containing M gene of influenza H3N2].
OBJECTIVE
To prepare the armored RNA containing M gene of influenza H3N2.


METHODS
The 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.


RESULTS
AR-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.


CONCLUSIONS
It 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.
Yu
Xin-fen
XF
Hangzhou Center for Disease Prevention and Control, Hangzhou 310006, China.
Pan
Jing-cao
JC
Huang
Zhi-cheng
ZC
Ye
Rong
R
Kou
Yu
Y
D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics*; D010957:Plasmids; D012367:RNA, Viral / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards; D014763:Viral Matrix Proteins / Q000235:genetics*
D012367:RNA, Viral; D014763:Viral Matrix Proteins