PrimerDesign-M was developed to meet the demands of primer design for highly variable sets of aligned sequences. It includes design requirements for next-generation sequences (NGS) including 454 sequences. It can also be used for primer and probe design for PCR, Sanger sequencing, and other systems with custom barcodes and DNA handles for universal primer annealing. The tool will design several alternative primer sets, whenever possible.
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PrimerDesign-M: a multiple-alignment based multiple-primer design tool for walking across variable genomes.
Yoon H, Leitner T.
Bioinformatics. 2014 Dec 17. pii: btu832.