Classification and review of free PCR primer design software

Guo, Jingwen; Starr, David S.; Guo, Huazhang

doi:10.1093/bioinformatics/btaa910

Cited by 20 publications

(16 citation statements)

References 46 publications

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“…Efficient PCR performance is highly dependent on PCR primer design, consequently one must spend a notable effort on the primer design. Well-designed PCR primers not only augment specificity and sensitivity but also reduce the effort spent on the experimental optimization 42 .…”

Section: Discussionmentioning

confidence: 99%

The validity evaluation of different 16srRNA gene primers for helicobacter detection urgently requesting to design new specific primers

Abdelmalek

Shokry

Hamed

et al. 2022

Sci Rep

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Molecular diagnosis of helicobacters by PCR is simpler, more accurate, and feasible compared to other diagnostic methods. Validity and accuracy are highly dependent on the PCR primer design, diffusion time, and mutation rate of helicobacters. This study aimed to design 16srRNA -specific primers for Helicobacter spp. and H. pylori. Application of comparative statistical analysis of the diagnostic utility of the most available 16srRNA genus-specific primers. The new primers were designed using bioinformatics tools (MAFFT MSA and Gblocks command line). A comparative study was applied on nine genus-specific 16srRNA primers in comparison to the ConsH using in silico and laboratory evaluation. The results demonstrated that the best specificity and sensitivity of the primers designed for this study compared to other primers. The comparative study revealed that the heminested outer/inner primers were the worst. Although H276, 16srRNA(a), HeliS/Heli-nest, and Hcom had acceptable diagnostic utility, false positive and false negative results were obtained. Specificity testing on clinical samples indicated a surprising result; that H. pylori was not the sole enemy that we were looking for, but the Non-Helicobacter pylori Helicobacters should be considered as a real risk prognostic for gastric diseases, consequently, a specific diagnosis and treatment should be developed. This study concluded that our designed primers were the most specific and sensitive in comparison with other primers. In addition, in silico evaluation is not accurate enough for primer assessment and that the laboratory evaluation is mandatory.

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Section: Discussionmentioning

confidence: 99%

The validity evaluation of different 16srRNA gene primers for helicobacter detection urgently requesting to design new specific primers

Abdelmalek

Shokry

Hamed

et al. 2022

Sci Rep

View full text Add to dashboard Cite

show abstract

Section: Discussionmentioning

confidence: 99%

Design of novel specific primers for helicobacter detection with diagnostic utility evaluation of different 16srRNA genus-specific primers: A comparative study

Abdelmalek

Shokry

Hamed

et al. 2021

Preprint

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Molecular diagnosis of helicobacters by PCR is simpler, more accurate, and feasible compared to other diagnostic methods. Validity and accuracy are highly dependent on the PCR primer design, diffusion time, and mutation rate of helicobacters. This study aimed to design 16srRNA -specific primers for Helicobacter spp. and H. pylori. Application of comparative statistical analysis of the diagnostic utility of the most available 16srRNA genus-specific primers. The new primers were designed using bioinformatics tools (MAFFT MSA and Gblocks command line). A comparative study was applied on nine genus-specific 16srRNA primers in comparison to the ConsH using Insilco and laboratory evaluation. The results demonstrated that the best specificity and sensitivity of the primers designed for this study compared to other primers. The comparative study revealed that the heminested outer/inner primers were the worst. Although H276,16srRNA(a), HeliS/Heli-nest, and Hcom had acceptable diagnostic utility, false positive and false negative results were obtained. Specificity testing on clinical samples indicated a surprising result; that H. pylori was not the sole enemy that we were looking for, but the NPH should be considered as a real risk prognostic for gastric diseases, consequently, a specific diagnosis and treatment should be developed. This study concluded that our designed primers were the most specific and sensitive in comparison with other primers. in addition, Insilco evaluation is not accurate enough for primer assessment and that the laboratory evaluation is mandatory.

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“…To identify strengths and weaknesses with rprimer, three other freely available tools (Gemi [ 10 ], DECIPHER [ 16 , 17 ], and openPrimeR [ 18 ]) were selected for comparison. Gemi was identified from a review of primer design programs published in 2020 [ 30 ]. It was selected because it is intended for similar purposes and uses the same overall strategy as rprimer (i.e.…”

Section: Case Study: Materials and Methodsmentioning

confidence: 99%

rprimer: an R/bioconductor package for design of degenerate oligos for sequence variable viruses

et al. 2022

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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.

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Classification and review of free PCR primer design software

Cited by 20 publications

References 46 publications

The validity evaluation of different 16srRNA gene primers for helicobacter detection urgently requesting to design new specific primers

The validity evaluation of different 16srRNA gene primers for helicobacter detection urgently requesting to design new specific primers

Design of novel specific primers for helicobacter detection with diagnostic utility evaluation of different 16srRNA genus-specific primers: A comparative study

rprimer: an R/bioconductor package for design of degenerate oligos for sequence variable viruses

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