「Unusual Features of the SARS-CoV-2 Genome Suggesting Sophisticated Laboratory Modification Rather Than Natural Evolution and Delineation of Its Probable Synthetic Route」
☝(おそらく「重症急性呼吸器症候群（SARS）」、（SARS-CoV1、SARS-CoV2、SARS-CoV3）、鳥インフルエンザウイルスマトリックス遺伝子（M300）、H5N1鳥の6つの遺伝子断片を含む2,248塩基の3V装甲L-RNAインフルエンザウイルス（HA300）— 2プラスミド共発現システムによって正常に発現され、装甲RNAのすべての特性を備えていることが実証されました。複数のウイルスアッセイのキャリブレーターとして3V装甲L-RNAを評価しました。 WHOInternational Standard for HCV RNA（NIBSC 96/790）を使用して、キメラ装甲L-RNAを較正し、10倍段階希釈で希釈して106〜102コピーを含むサンプルを取得しました。結論として、装甲L-RNA調製に使用したアプローチは実用的であり、マルチプレックスRNAウイルスアッセイの品質管理の労力とコストを削減できます。さらに、定量的検出のための国際単位をキメラ装甲RNAに割り当てることができます。
［榮語原文］
RNase-resistant, noninfectious virus-like particles containing exogenous RNA sequences (armored RNA) are good candidates as RNA controls and standards in RNA virus detection. However, the length of RNA packaged in the virus-like particles with high efficiency is usually less than 500 bases. In this study, we describe a method for producing armored L-RNA. Armored L-RNA is a complex of MS2 bacteriophage coat protein and RNA produced in Escherichia coli by the induction of a two-plasmid coexpression system in which the coat protein and maturase are expressed from one plasmid and the target RNA sequence with modified MS2 stem-loop (pac site) is transcribed from another plasmid. A 3V armored L-RNA of 2,248 bases containing six gene fragments— hepatitis C virus, severe acute respiratory syndrome coronavirus (SARS-CoV1, SARS-CoV2, and SARS-CoV3), avian influenza virus matrix gene (M300), and H5N1 avian influenza virus (HA300)—was successfully expressed by the two-plasmid coexpression system and was demonstrated to have all of the characteristics of armored RNA. We evaluated the 3V armored L-RNA as a calibrator for multiple virus assays. We used the WHOInternational Standard for HCV RNA (NIBSC 96/790) to calibrate the chimeric armored L-RNA, which was diluted by 10-fold serial dilutions to obtain samples containing 106 to 102 copies. In conclusion, the approach we used for armored L-RNA preparation is practical and could reduce the labor and cost of quality control in multiplex RNA virus assays. Furthermore, we can assign the chimeric armored RNA with an international unit for quantitative detection.
From the off-guardian, January 3, 2021, “What Vaccine Trials?” by Iain Davis:
“…the WHO protocols Pfizer used to produce the mRNA [for the vaccine] do not appear to identify any nucleotide sequences that are unique to the SARS-CoV-2 virus. When investigator Fran Leader questioned Pfizer they confirmed: ‘The DNA template does not come directly from an isolated virus from an infected person’.”
