Preparation and Evaluation of Ribonuclease-Resistant Viral HIV RNA Standards Based on Armored RNA Technology

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GHOLAMI MOHAMMAD; RAVANSHAD MEHRDAD; BAESI KAZEM; Samiee Siamak M.; Hosseini Rozbahani Negin; MOHRAZ MINOO

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Journal Paper

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Journal: Iranian Biomedical Journal Year:2018 | Volume:22 | Issue:6 Page(s): 394-400

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Title

Preparation and Evaluation of Ribonuclease-Resistant Viral HIV RNA Standards Based on Armored RNA Technology

Author(s)

Keywords

HIV

Real

time PCR

Virus

like particle

Abstract

Background: The human immunodeficiency Virus type 1 (HIV-1) is an infectious viral agent that gradually extinguishes the immune system, resulting in acquired immune deficiency syndrome (AIDS). The aim of this study was to construct an RNA-positive control based on armored (AR) RNA technology, using HIV-1 RNA as a model. Methods: The MS2 maturase, a coat protein gene (at positions 1765 to 1787) and HIV-1 pol gene were cloned into pET-32a plasmid. The prepared plasmid was transformed into Escherichia coli strain BL2 (DE3), and the expression of the construct was induced by 1 mM of isopropyl-L-thio-D-galactopyranoside (IPTG) at 37 ° C for 16 h to obtain the fabricated AR RNA. The AR RNA was precipitated and purified using polyethylene glycol and Sephacryl S-200 chromatography. Results: The stability of AR RNA was evaluated by treatment with DNase I and RNase A and confirmed by transmission electron microscopy and gel agarose electrophoresis. Tenfold serial dilution of AR RNA from 101 to 105 was prepared. Real-time PCR assays had a range of detection between 101 and 105. In addition, R2 value was 0. 998, and the slope of the standard curve was-3. 33. Conclusion: Prepared AR RNA, as a positive control, could be used as a basis for launching an in-house HIV-1 Virus assay and other infectious agents. It can be readily available to laboratories and HIV research centers. The AR RNA is non-infectious and highly resistant to ribonuclease enzyme and can reduce the risk of infection in the clinical laboratory.

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APA: Copy

GHOLAMI, MOHAMMAD, RAVANSHAD, MEHRDAD, BAESI, KAZEM, Samiee, Siamak M., Hosseini Rozbahani, Negin, & MOHRAZ, MINOO. (2018). Preparation and Evaluation of Ribonuclease-Resistant Viral HIV RNA Standards Based on Armored RNA Technology. IRANIAN BIOMEDICAL JOURNAL, 22(6), 394-400. SID. https://sid.ir/paper/746583/en

Vancouver: Copy

GHOLAMI MOHAMMAD, RAVANSHAD MEHRDAD, BAESI KAZEM, Samiee Siamak M., Hosseini Rozbahani Negin, MOHRAZ MINOO. Preparation and Evaluation of Ribonuclease-Resistant Viral HIV RNA Standards Based on Armored RNA Technology. IRANIAN BIOMEDICAL JOURNAL[Internet]. 2018;22(6):394-400. Available from: https://sid.ir/paper/746583/en

IEEE: Copy

MOHAMMAD GHOLAMI, MEHRDAD RAVANSHAD, KAZEM BAESI, Siamak M. Samiee, Negin Hosseini Rozbahani, and MINOO MOHRAZ, “Preparation and Evaluation of Ribonuclease-Resistant Viral HIV RNA Standards Based on Armored RNA Technology,” IRANIAN BIOMEDICAL JOURNAL, vol. 22, no. 6, pp. 394–400, 2018, [Online]. Available: https://sid.ir/paper/746583/en

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