Computational protocol: One-step real-time RT-PCR assays for serotyping dengue virus in clinical samples

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Protocol publication

[…] To be able to find serotype-specific conserved regions, all whole genome sequences of DENV1-4 available at the design stage (2014–10–31) were downloaded from NCBI and used for assay design. Multiple sequence alignments containing the genomic sequences of the individual serotypes and alignments containing all DENV genomic sequences were created using CLC Genomics Workbench 7.5 (www.clcbio.com). Primers and probes were constructed using in-house software. Melting temperatures (Tm) were verified using Primer Express® v3.0 (Applied Biosystems®). Theoretical specificity of the systems was investigated using BLAST against the NCBI nucleotide database with very loose match criteria (word-size = 7, E-cutoff = 1000, match/mismatch cost +1/-1, gap cost 5/2). […]

Pipeline specifications

Software tools CLC Genomics Workbench, CLC Assembly Cell, Primer Express
Applications WGS analysis, qPCR
Organisms Homo sapiens
Chemicals Nucleotides