Short-Amplicon Strategy
Definition
A PCR design that targets the shortest possible fragment length: often 80-150 bp: within a diagnostic locus such as COI. Shorter targets are more likely to be intact in degraded templates.
- Typical fragment size
- About 80 to 150 base pairs
- Target locus example
- COI (cytochrome c oxidase subunit I)
- Rationale
- Shorter fragments are more likely to survive DNA degradation intact
- Common use case
- Processed or degraded wildlife products such as powders, leather, or cooked meat
Common questions
Why not just use the standard full-length COI barcode region for degraded samples?+
The standard barcode region, roughly 650 base pairs, is often too long to survive intact once DNA has been fragmented by heat, processing, or time, so amplification fails outright on many degraded exhibits. A short-amplicon target within the same gene is far more likely to amplify from the same sample.
Does using a shorter amplicon reduce species identification confidence?+
It can, because a shorter sequence carries less discriminating information than the full barcode, so species assignment sometimes has lower resolution or cannot distinguish closely related species that differ only outside the short target region. Analysts weigh this trade-off against the alternative of no result at all.
Related terms
- Collagen Peptide Barcoding
- Species identification from trypsin-digested collagen analysed by mass spectrometry. Peptide sequences differ between species at diagnostic positions that survive tanning and heat...
- DNA Fragmentation
- Physical breakage of the DNA backbone into short pieces. In degraded samples the average fragment length may be 50-200 bp, making long...
- Library Preparation (aDNA)
- Enzymatic end-repair, adapter ligation, and amplification of all DNA fragments in a sample regardless of length, used in ancient-DNA and degraded-sample workflows....
- Metagenomics
- The direct sequencing of all DNA extracted from an environmental sample, bypassing the need to culture individual organisms. It produces a community-wide...
- PCR Inhibitor
- A co-extracted substance that interferes with Taq polymerase activity or primer annealing. Common inhibitors in forensic samples include haemoglobin (from blood on...