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
This record last updated: 12-13-2012