Calibration of the real-time RT-PCR assay for HCV, HIV-1, SARS-CoV1, and SARS-CoV2 02996 article

Preparation of a Chimeric Armored RNA as a Versatile Calibrator for Multiple Virus Assays
Qiuying Huang, Yangjian Cheng, Qiwei Guo, Qingge Li
Clinical Chemistry, Volume 52, Issue 7, 1 July 2006, Pages 1446–1448, https://doi.org/10.1373/clinchem.2006.069971
Published:
01 July 2006 We used a pure RNA transcript fragment of SARS-CoV2 (BNI) to calibrate the chimeric armored RNA, then used the chimeric armored RNA to prepare calibrators of the 4 real-time reverse transcription-PCR (RT-PCR) assays (Fig. 11 ; also see Fig. 1 in the online Data Supplement) based on displacing probes (11). The linear range for each assay did not change when the calibrators were stored at 37 °C for 2 weeks, at 4 °C for 6 months, or at −20 °C for 1 year.
Our work indicates that multiple target sequences can be encapsulated into a single armored RNA species to serve as a common calibrator for detection of different RNA viruses. Chimeric armored RNA of even larger size may be prepared similarly, as indicated by our finding that by deleting some disposable sequences between the multiple cloning site and the transcription terminator, we were able to increase packaging capacity of the pAR-1 vector without affecting packaging efficiency (data not shown). Thus, the chimeric, multitarget approach for armored RNA preparation is practical and could reduce the labor and cost for quality control of multiplex RNA virus assays.
Figure 1.
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Calibration of the real-time RT-PCR assay for HCV, HIV-1, SARS-CoV1, and SARS-CoV2.

We used a pure RNA transcript fragment of SARS-CoV2 (BNI) to calibrate the chimeric armored RNA, then used the chimeric armored RNA to prepare calibrators of the 4 real-time reverse transcription-PCR (RT-PCR) assays (Fig. 11 ; also see Fig. 1 in the online Data Supplement) based on displacing probes (11). The linear range for each assay did not change when the calibrators were stored at 37 °C for 2 weeks, at 4 °C for 6 months, or at −20 °C for 1 year.
Our work indicates that multiple target sequences can be encapsulated into a single armored RNA species to serve as a common calibrator for detection of different RNA viruses. Chimeric armored RNA of even larger size may be prepared similarly, as indicated by our finding that by deleting some disposable sequences between the multiple cloning site and the transcription terminator, we were able to increase packaging capacity of the pAR-1 vector without affecting packaging efficiency (data not shown). Thus, the chimeric, multitarget approach for armored RNA preparation is practical and could reduce the labor and cost for quality control of multiplex RNA virus assays.
Figure 1.
Open in new tabDownload slide
Calibration of the real-time RT-PCR assay for HCV, HIV-1, SARS-CoV1, and SARS-CoV2.