RNA: packaged and protected by VLPs

Fang, Po-Yu; Bowman, Jessica C.; Ramos, Lizzette M. Gómez; Hsiao, Chiaolong; Williams, Loren Dean

doi:10.1039/c8ra02084a

Cited by 19 publications

(34 citation statements)

References 75 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The two-plasmid system has been used successfully in previous work for in vivo reconstitution of Qβ around target RNAs. 30 , 42 …”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, the presence of RBS upstream of target RNA has been shown to reduce packaging efficiency of RNAs into VLPs due to competitive binding of ribosomes versus Qβ CPs to the target RNA. 30 Therefore, for the one- and two-plasmid systems, the RBS was removed upstream of the SDM, but RBS was retained upstream of the Qβ CPs to enable protein translation. In vitro transcription of SDM RNAs yielded approximately 150 μg of SDM RNAs per reaction; denaturing urea polyacrylamide gel electrophoresis of the transcribed SDM RNA confirmed the ∼622 nt RNA product ( Figure S3A ).…”

Section: Resultsmentioning

confidence: 99%

“…To achieve efficient in vivo reconstitution, careful design considerations need to be taken in account, such as length of the target RNA and its molecular features, i.e ., deletion of ribosome binding sites (for enhanced safety) and appendage of Qβ hairpin (for encapsidation). 30 Here, we developed such methods for the Qβ VLP system. The structural integrity and stability of the resulting nanoparticle positive controls were assessed over time using a combination of transmission electron microscopy (TEM), size exclusion chromatography (SEC), dynamic light scattering (DLS), and electrophoretic mobility assays.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

Chan

Ignacio

et al. 2020

ACS Nano

View full text Add to dashboard Cite

Coronavirus disease 2019 (COVID-19) is a highly transmissible disease that has affected more than 90% of the countries worldwide. At least 17 million individuals have been infected, and some countries are still battling first or second waves of the pandemic. Nucleic acid tests, especially reverse transcription polymerase chain reaction (RT-PCR), have become the workhorse for early detection of COVID-19 infection. Positive controls for the molecular assays have been developed to validate each test and to provide high accuracy. However, most available positive controls require cold-chain distribution and cannot serve as full-process control. To overcome these shortcomings, we report the production of biomimetic virus-like particles (VLPs) as SARS-CoV-2 positive controls. A SARS-CoV-2 detection module for RT-PCR was encapsidated into VLPs from a bacteriophage and a plant virus. The chimeric VLPs were obtained either by in vivo reconstitution and coexpression of the target detection module and coat proteins or by in vitro assembly of purified detection module RNA sequences and coat proteins. These VLP-based positive controls mimic SARS-CoV-2 packaged ribonucleic acid (RNA) while being noninfectious. Most importantly, we demonstrated that the positive controls are scalable, stable, and can serve broadly as controls, from RNA extraction to PCR in clinical settings.

show abstract

“…The two-plasmid system has been used successfully in previous work for in vivo reconstitution of Qβ around target RNAs. 30 , 42 …”

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

Chan

Ignacio

et al. 2020

ACS Nano

View full text Add to dashboard Cite

show abstract

“…The two-plasmid system has been used successfully in previous work for in vivo reconstitution of Qβ around target RNAs. 30,42 produced from a one-plasmid system, where the gene of the Qβ CPs and SDM RNAs were cloned in one vector (pCDFDuet™-1). Qβ 2P-C19 VLPs were produced from a two-plasmid system, where Qβ CP was derived from pCDFDuet™-Qβ and SDM RNA was produced from pET-28a (+).…”

Section: Design Of the Sars-cov-2 Detection Module (Sdm)mentioning

confidence: 99%

“…deletion of ribosome binding sites (for enhanced safety) and appendage of Qβ hairpin (for encapsidation). 30 Here we developed such methods for the Qβ VLP system. The structural integrity and stability of the resulting nanoparticle positive controls were assessed over time using a combination of transmission electron microscopy (TEM), size exclusion chromatography (SEC), dynamic light scattering (DLS) and electrophoretic mobility assays.…”

mentioning

confidence: 99%

Biomimetic Virus-like Particles as SARS-CoV-2 Positive Controls for RT-PCR Diagnostics

Chan

Ignacio

Metha

et al. 2020

Preprint

View full text Add to dashboard Cite

Coronavirus disease 2019 (COVID-19) is a highly transmissible disease that has affected more than 90% of the countries worldwide. At least 17 million individuals have been infected, and some countries are still battling first or second waves of the pandemic. Nucleic acid tests, especially reverse-transcription polymerase chain reaction (RT-PCR), have become the workhorse for early detection of COVID-19 infection. Positive controls for the molecular assays have been developed to validate each test and to provide high accuracy. However, most available positive controls require cold-chain distribution and cannot serve as full-process control. To overcome these shortcomings, we report the production of biomimetic virus-like particles (VLPs) as SARS-CoV-2 positive controls. A SARS-CoV-2 detection module for RT-PCR was encapsidated into VLPs from a bacteriophage and a plant virus. The chimeric VLPs were obtained either by in vivo reconstitution and co-expression of the target detection module and coat proteins or by in vitro assembly of purified detection module RNA sequences and coat proteins. These VLP-based positive controls mimic SARS-CoV-2 packaged RNA while being non-infectious. Most importantly, we demonstrated that the positive controls are scalable, stable, and can serve broadly as controls, from RNA extraction to PCR in clinical settings.

show abstract

Changes in Structure, Dispersity and Phase Behavior of Proteins: The Virus-Like Particles in the Presence of Nucleic Acids

Pusara,

Valentic,

Wenzel

et al. 2024

Dispersity, Structure and Phase Changes of Proteins and Bio Agglomerates in Biotechnological Processes

View full text Add to dashboard Cite

RNA: packaged and protected by VLPs

Abstract: VLP packaging is most efficient for compact RNA, and protects RNA against assault by small diffusible damaging agents.

Cited by 19 publications

References 75 publications

Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

Biomimetic Virus-like Particles as SARS-CoV-2 Positive Controls for RT-PCR Diagnostics

Changes in Structure, Dispersity and Phase Behavior of Proteins: The Virus-Like Particles in the Presence of Nucleic Acids

Contact Info

Product

Resources

About