SPADA Draft Documents

Multiplex Primer Dimer Check

Check for all possible primer dimer interactions Compute cross-hybridizations of all primer candidates against all amplicons Compute false amplicons of all primers against background database in silico analysis and combinations of singleplexes Capture singleplex performance metrics Tools that allow for replacement of poor performing primers Tools that allow for replacement of poor performing primers in silico analysis and combinations of singleplexes Capture multiplexplex performance metrics Tools for analyzing existing assays using new target variants Software summary of primer/probe performance Analyze input and performance metrics to determine best practices

input for multiplex algorithm

Multiplex Primer-amplicon Check

Compute cross-hybridizations of all primer candidates against all amplicons Multiplex algorithm needs to minimize false amplicons to allow even amplification of all targets Determine optimum combination of singplexes that are mutually compatible with eachother Capture singleplex performance, link to input file information, Allow for future A.I. meta-analysis Hold constant primers that work, redesign primers for targets that don't work Hold constant primers that work, redesign primers for targets that don't work Determine optimum combination of singplexes that are mutually compatible with eachother Capture multiplex performance, link to input file information, Allow for future A.I. meta-analysis Ensure that existing assays work on new target variants Summary of final design: thermodynamic, heuristic scores, specificity, sequence alignment validate and verify predefined standards for traceability, accuracy, reliability, and precision

Multiplex Primer-background Check

Multiplexing

Experimental Validation Re-design

Determine Performance of Singeplexes Redesign of failed singleplexes

Redesign of failed mutliplexes

Multiplexing

Experimental Validation

Determine Performance of Multiplex

Post-design analysis

Assay Stewardship

Final Report

Metrology

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