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Oligoanalyzer online software

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The OligoAnalyzer is an online software tool provided by Integrated DNA Technologies. It allows users to analyze the properties of DNA or RNA oligonucleotide sequences, including melting temperature, self-complementarity, and potential for secondary structures.

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OligoAnalyzer (26 citations) OligoAnalyzer software (6 citations)

4 protocols using oligoanalyzer online software

1

SARS-CoV-2 Primer and Probe Design Validation

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Primer and probe candidates were designed for each CS using Primer3 (Untergasser et al. 2012 (link)). The specificities of the primers and probes were assessed by Primer-BLAST (Ye et al. 2012 (link)) comparisons against the complete RefSeq RNA databases for Homo sapiens (taxid: 9606), bacteria (taxid: 2), fungi (taxid: 4751), and Apicomplexa (taxid: 5794). OligoAnalyzer online software (Integrated DNA Technologies, Coralville, IA, USA) was used to check for primer dimers and secondary structure formation.
The EMBOSS ‘PrimerSearch’ program (Rice et al. 2000 (link)) was used to check primer pairs against the three SARS-CoV-2 genome datasets used in this study, with 0%, 5%, and 10% mismatches between the primers and templates allowed. For comparison, nucleotide sequences from 16 primer sets that are either used widely worldwide or have been developed by domestic researchers were included in the in silico analysis (Corman et al. 2020 (link); Won et al. 2020 (link)).
Jeong H., Lee S., Ko J., Ko M, & Seo H.W. (2022). Identification of conserved regions from 230,163 SARS-CoV-2 genomes and their use in diagnostic PCR primer design. Genes & Genomics, 44(8), 899-912.
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2

Designing Gonococcal LAMP Primers

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A set of special LAMP primers, targeting on the orf1 gene of N. gonorrhoeae (GenBank Accession No. M84113), was designed according to the LAMP principle. The LAMP primers consist of two outer primers (F3 and F3), two inner primers (FIP and BIP), and two loop primers (LF and LB). The specificity of N. gonorrhoeae-LAMP primers was analyzed using the Basic Local Alignment Search Tool (BLAST). Moreover, OligoAnalyzer online software (V3.1; Integrated DNA Technologies, Coralville, IA, United States) was employed for primer, dimer, and secondary structure investigation. The details of N. gonorrhoeae-LAMP primers’ location and sequences are shown in Figure 2 and Table 1, respectively.
Chen X., Zhou Q., Wu X., Wang S., Liu R., Dong S, & Yuan W. (2021). Visual and Rapid Diagnosis of Neisseria gonorrhoeae Using Loop-Mediated Isothermal Amplification Combined With a Polymer Nanoparticle–Based Biosensor in Clinical Application. Frontiers in Molecular Biosciences, 8, 702134.
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3

SARS-CoV-2 Detection via RT-MCDA

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Two sets of RT-MCDA primers (F1ab-MCDA and N-MCDA primer sets), which targeted the F1ab gene and N gene, respectively, of SARS-CoV-2 (GenBank MN908947, Wuhan-Hu-1), were designed according the MCDA principle (figure 3). Each MCDA primer set consists of two displacement primers (F1 and F2), two cross-primers (CP1 and CP2), and six amplification primers (C1, D1, R1, C2, D2 and R2). The specificity of the F1ab- and N-MCDA primer sets were analysed using the National Center for Biotechnology Information basic local alignment search tool. Moreover, OligoAnalyzer online software (V3.1; Integrated DNA Technologies, Coralville, IA, USA) was employed for primer, dimer and secondary structure investigation. The details of RT-MCDA primer design, primer location, sequences and modifications are listed in figure 3 and supplementary table S1. All oligomers were synthesised and purified by Tianyi-Huiyuan Biotech (Beijing, China) at HPLC purification grade.
Li S., Jiang W., Huang J., Liu Y., Ren L., Zhuang L., Zheng Q., Wang M., Yang R., Zeng Y, & Wang Y. (2020). Highly sensitive and specific diagnosis of COVID-19 by reverse transcription multiple cross-displacement amplification-labelled nanoparticles biosensor. The European Respiratory Journal, 56(6), 2002060.
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4

Nested Primer Design for MCDA

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Based on MCDA reaction mechanisms, five pairs of primers based on orf1 (GenBank Accession No. M84113) were designed using primer Explorer V5 and PRIMER PREMIER 5.0 software (Eiken Chemical, Japan). A set of N. gonorrhoeae-MCDA primers, including displacement primers (F1 and F2), cross primers (CP1 and CP2), and amplification primers (D1, D2, C1, C2, R1, and R2), was also generated (Table 1). N. gonorrhoeae-MCDA primer specificity was verified using the BLAST analysis tool. In addition, Oligo Analyzer online software (V3.1; Integrated DNA Technologies, Coralville, IA, United States) was used for primer secondary structure and dimer investigations. MCDA primer sequences are shown (Table 1). All primers were synthesized and purified at TsingKe Biotech Co., Ltd. (Beijing, China) using high performance liquid chromatography purification grade.
Chen X., Huang L., Zhou Q., Tan Y., Tan X, & Dong S. (2021). A Nanoparticle-Based Biosensor Combined With Multiple Cross Displacement Amplification for the Rapid and Visual Diagnosis of Neisseria gonorrhoeae in Clinical Application. Frontiers in Microbiology, 12, 747140.
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