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We are analyzing https://bmcbioinformatics.biomedcentral.com/articles/10.1186/s12859-022-04781-0.

Title:
rprimer: an R/bioconductor package for design of degenerate oligos for sequence variable viruses | BMC Bioinformatics | Full Text
Description:
Background This paper presents a new R/Bioconductor package, rprimer, for design of degenerate oligos and PCR assays for sequence variable viruses. A multiple DNA sequence alignment is used as input data, while the outputs consist of comprehensive tables (data frames) and dashboard-like plots. The workflow can be run directly from the R console or through a graphical user interface (Shiny application). Here, rprimer is demonstrated and evaluated by using it to design two norovirus genogroup I (GI) assays: one RT-qPCR assay for quantitative detection and one RT‑PCR assay for Sanger sequencing and polymerase-capsid based genotyping. Results The assays generated were evaluated using stool samples testing positive for norovirus GI. The RT-qPCR assay accurately amplified and quantified all samples and showed comparable performance to a widely-used standardised assay, while the RT-PCR assay resulted in successful sequencing and genotyping of all samples. Merits and limitations of the package were identified through comparison with three similar freely available software packages. Several features were comparable across the different tools, but important advantages of rprimer were its speed, flexibility in oligo design and capacity for visualisation. Conclusions An R/Bioconductor package, rprimer, was developed and shown to be successful in designing primers and probes for quantitative detection and genotyping of a sequence-variable virus. The package provides an efficient, flexible and visual approach to degenerate oligo design, and can therefore assist in virus research and method development.
Website Age:
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Keywords {🔍}

assay, design, sequence, primers, primer, sequences, assays, oligo, rprimer, article, target, package, norovirus, google, scholar, oligos, alignment, degenerate, pcr, data, rtqpcr, software, fig, dna, probes, consensus, variable, cas, rtpcr, degeneracy, input, decipher, openprimer, binding, region, reference, user, reverse, part, variants, intended, study, bmc, bases, information, viruses, genotyping, step, match, process,

Topics {✒️}

springer nature real-time reverse transcription-pcr multiplex real-time rt-pcr implementation overview digital pcr sharing discussion case study redesigned duplex rt-qpcr project home page real-time rt-pcr rna-dependent rna polymerase food chain—horizontal method polymerase-capsid based genotyping polymerase-capsid-based genotyping conventional rt-pcr assay data import page bmc infect dis author information authors step rt-pcr system rt-pcr assay resulted polymerase chain reaction decipher recommend/require external case study sequence-variable virus difficult primer-binding sites rt-qpcr assay target privacy choices/manage cookies false-positive signals compared widely-adopted data structures real-time pcr top-scoring forward primer degenerate pcr primers additional information publisher' pcr primer design rt-pcr products state privacy rights rt-qpcr experiment step rt-pcr full-length genomes complete minimum information 25 µl reaction mix synthesised pcr products sequence variable viruses rt‑pcr assay rt-pcr assay sequence variable targets taq dna polymerase pcr primers prediction step rt-qpcr rt-pcr applications

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         description:This paper presents a new R/Bioconductor package, rprimer, for design of degenerate oligos and PCR assays for sequence variable viruses. A multiple DNA sequence alignment is used as input data, while the outputs consist of comprehensive tables (data frames) and dashboard-like plots. The workflow can be run directly from the R console or through a graphical user interface (Shiny application). Here, rprimer is demonstrated and evaluated by using it to design two norovirus genogroup I (GI) assays: one RT-qPCR assay for quantitative detection and one RT‑PCR assay for Sanger sequencing and polymerase-capsid based genotyping. The assays generated were evaluated using stool samples testing positive for norovirus GI. The RT-qPCR assay accurately amplified and quantified all samples and showed comparable performance to a widely-used standardised assay, while the RT-PCR assay resulted in successful sequencing and genotyping of all samples. Merits and limitations of the package were identified through comparison with three similar freely available software packages. Several features were comparable across the different tools, but important advantages of rprimer were its speed, flexibility in oligo design and capacity for visualisation. An R/Bioconductor package, rprimer, was developed and shown to be successful in designing primers and probes for quantitative detection and genotyping of a sequence-variable virus. The package provides an efficient, flexible and visual approach to degenerate oligo design, and can therefore assist in virus research and method development.
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      headline:rprimer: an R/bioconductor package for design of degenerate oligos for sequence variable viruses
      description:This paper presents a new R/Bioconductor package, rprimer, for design of degenerate oligos and PCR assays for sequence variable viruses. A multiple DNA sequence alignment is used as input data, while the outputs consist of comprehensive tables (data frames) and dashboard-like plots. The workflow can be run directly from the R console or through a graphical user interface (Shiny application). Here, rprimer is demonstrated and evaluated by using it to design two norovirus genogroup I (GI) assays: one RT-qPCR assay for quantitative detection and one RT‑PCR assay for Sanger sequencing and polymerase-capsid based genotyping. The assays generated were evaluated using stool samples testing positive for norovirus GI. The RT-qPCR assay accurately amplified and quantified all samples and showed comparable performance to a widely-used standardised assay, while the RT-PCR assay resulted in successful sequencing and genotyping of all samples. Merits and limitations of the package were identified through comparison with three similar freely available software packages. Several features were comparable across the different tools, but important advantages of rprimer were its speed, flexibility in oligo design and capacity for visualisation. An R/Bioconductor package, rprimer, was developed and shown to be successful in designing primers and probes for quantitative detection and genotyping of a sequence-variable virus. The package provides an efficient, flexible and visual approach to degenerate oligo design, and can therefore assist in virus research and method development.
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      dateModified:2022-06-18T00:00:00Z
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         RT-PCR
         qPCR
         Digital PCR
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         Genotyping
         Bioinformatics
         Microarrays
         Computational Biology/Bioinformatics
         Computer Appl. in Life Sciences
         Algorithms
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      name:Department of Medical Sciences, Zoonosis Science Centre, Uppsala University, Uppsala, Sweden
      name:Section of Clinical Virology, Department of Medical Sciences, Uppsala University, Uppsala, Sweden
      name:European Union Reference Laboratory for Foodborne Viruses, Swedish Food Agency, Uppsala, Sweden
      name:Department of Medical Sciences, Zoonosis Science Centre, Uppsala University, Uppsala, Sweden

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