Create Exon Junction Primers

What you can do:
Automated pipeline for RT-PCR primer design, targeted at exon-junction sites.
Highlights:
  • Supported organism are human, mouse, rat and fruitfly.
  • This tool specifically tries to design Realtime-primers overlapping exon-exon junctions.
  • The resulting amplicons should only amplify cDNA sequences since these junctions are not present in unspliced genomic sequences.
  • The sequences are fetched from UCSC databases, based on regions corresponding to the supplied RefSeq Symbol.
  • Next, primers will be tested on each of the exon-exon boundaries to check if they form primer dimers and hairpin structures, have an acceptable GC-content and melting temperature.
  • Acceptable primers are then combined to form amplicons.
  • Amplicons of acceptable size are then folded used mfold to check for secondary structures and blasted against the refseq_RNA database to check specificity.
  • A short summary is provided listing the primers, the amplicon and all targets (multiple targets are allowed if they are isoforms of the same gene).
Keywords:
  • PCR primers
  • oligos databases
  • design tools
  • primer design
Literature & Tutorials:
PubMed Link:
This record last updated: 12-13-2012
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