About JAK2
The JAK2 gene (Janus Kinase 2) on chromosome 9p24 The V617F mutation in exon 14 is found in>95% of polycythemia vera and ~50% of essential thrombocythemia and primary myelofibrosis. Key targets include Exons 12–24 (JH2 pseudokinase, JH1 kinase) for comprehensive mutation screening.
Mutations: V617F, exon 12 mutations (N542-E543del) | GC content: 50% | Target exons: Exons 12–24 (JH2 pseudokinase, JH1 kinase)
JAK2 Primer Design Challenges
- V617F dominance: Valine 617 in exon 14 is the single most important mutation site, but its high GC context requires optimized amplification
- Exon 12 mutations: N542-E543del and other in-frame indels in the pseudokinase domain require additional amplicon coverage
- Moderate GC (50%): Balanced GC content allows standard PCR, but the JH2 pseudokinase domain (exon 12–14) has localized higher-GC stretches
Recommended Primer Design Parameters for JAK2
| Parameter | Standard Exons | Optimized Regions |
|---|---|---|
| Primer length | 20–22 nt | 22–25 nt |
| GC content | 45–55% | 50–60% |
| Tm | 58–62°C | 60–65°C |
| Amplicon size | 150–300 bp | 180–350 bp |
| Annealing temp | 58–60°C | 60–64°C (touchdown) |
| PCR additive | Standard | Optional 3–5% DMSO for problematic regions |
Recommended Primer Sequences for JAK2
| Target Region | Forward Primer (5′→3′) | Reverse Primer (5′→3′) | Amplicon |
|---|---|---|---|
| Exon 14 (V617F) | 5′-TGCTCTGAGACGTTGAGTCA-3′ | 5′-GCTGTGATCCTGAAACTGGA-3′ | 186 bp |
| Exon 12 (JH2 region) | 5′-ACTCAGCAGGATGAAGCCAA-3′ | 5′-CAGGCCAAGATCTTCTTCGT-3′ | 175 bp |
Key SNPs to Avoid in Primer Binding Sites
When designing JAK2 primers, avoid these clinically significant variants:
- rs77375493 (V617F) — Exon 14 JAK-STAT activating mutation in MPN
- JAK2 exon 12 (N542-E543del) — Exon 12 in-frame deletion in PV
Clinical Validation Required
All JAK2 primers designed with VigyanLLM are for research use only. Clinical diagnostic applications require additional wet-lab validation, Sanger sequencing confirmation, and regulatory approval before patient use.
Design JAK2 Primers Now
Pre-configured with JAK2-specific parameters. Use our primer design, Tm calculator, and GC content tools for optimal results.