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RT-qPCR · Exon-aware · Hydrolysis probe

RT-qPCR Primer Design Tool

Design and prioritize RT-qPCR primer pairs with exon-aware ranking, genomic-DNA risk assessment and optional hydrolysis-probe candidates.

Start RT-qPCR + Probe Design Open Sequence-only Primer Design

RefSeq workflow: start from a gene + organism, choose the transcript, then load its sequence. Sequence-only workflow: use Primer Design when you already have a DNA template sequence.

Primer3-based design Exon-aware ranking gDNA-risk review Hydrolysis-probe candidates
Exon-aware RT-qPCR design cDNA Exon 3 Exon 4 Forward primer across junction Reverse primer Genomic DNA Intron The same exon junction is not contiguous in genomic DNA
Exon-junction or inter-exon designs can help reduce genomic-DNA interference when the transcript structure supports them.

Hydrolysis Probe Candidates

Primer–probe arrangement Forward primer Probe Reverse primer The probe lies inside the amplicon and is evaluated separately from the primer pair.
Probe candidates are sequence-level suggestions only; fluorophore, quencher and final chemistry are not selected by the tool.
Important. Probe sequence quality does not guarantee assay performance. Probe labeling chemistry, template abundance, reaction composition and instrument settings can all affect the final assay.
Research use: exon-aware ranking and probe candidates require experimental specificity, efficiency and control validation.

Related Molecular Design Lab

Design Assay Primer Design Tool Standard PCR Primer Design Peptide Analysis MHC / HLA Prediction Projects