design

Design

RealTimeDesign Software


Good probe and primer design is at the heart of any successful real-time PCR assay. RealTimeDesign™ software is an easy yet powerful application, and is free upon registration. The program uses sophisticated algorithms to select optimal probe and primer sequences for your target of interest based on BHQ and BHQplus probe-types. With RealTimeDesign Software, you may design oligo sequences for qPCR including gene expression analysis, SNP genotyping, and multiplexed assay designs.

RealTimeDesign software
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Fast Design in Express Mode


You can design an optimal assay by utilizing RealTimeDesign software’s express mode. Simply follow these steps:

  1. From the Main Menu select “Design Genomic Assay”
  2. Enter your target sequence region of interest up to 1000 bp, or enter the GenBank accession number Multiple Sequences can be indicated by including a “>Gene Name” in the line above each sequence for example:
    • >Gene Name A
    • Sequence A
    • >Gene Name B
    • Sequence B
  3. Select your Application, whether qPCR or SNP genotyping, and the intended probe format. Indicate if this is a single-plex or multiplexed assay.
  4. If this is a SNP assay, annotate the location of the SNP in brackets, with each allele separated by a slash (Examples: [A/T] or [G/T]). This will let the software know to design two probes against each allele at that position.
  5. For gene expression analysis you can specify an exon-exon junction (intron splice site) by including a ~ in the sequence. This will modify the software settings to propose an assay spanning the ~ feature in the sequence. This strategy is often useful to minimize amplification of genomic DNA in a assay targeting RNA.
  6. Click Next and select your instrument for the recommended dye options. Select the design parameter sets, and then click next again to start design.
  7. Review results! You can also download the oligo designs or add them to the shopping cart for synthesis.


real-time PCR probe design