About FBXW7
The FBXW7 gene (F-Box and WD Repeat Domain Containing 7) on chromosome 4q31 Inactivating missense mutations at arginine residues 465, 479, and 505 in the WD40 domain occur in T-ALL, colorectal, and gastric cancers. Key targets include Exons 2–10 (WD40 repeats in exons 4–10) for comprehensive mutation screening.
Mutations: R465C, R465H, R479Q, R505L | GC content: 46% | Target exons: Exons 2–10 (WD40 repeats in exons 4–10)
FBXW7 Primer Design Challenges
- WD40 domain hotspot: Arginine 465 mutations in exon 8 affect substrate binding and require focused amplicon design
- Low GC content (46%): AT-rich introns flanking exons 3–6 require lower annealing temperatures and longer primer design
- Multiple target substrates: FBXW7’s role in degrading multiple oncoproteins requires complete mutation coverage across all WD40 repeats
Recommended Primer Design Parameters for FBXW7
| Parameter | Standard Exons | Low-GC / AT-Rich 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 | Longer primers (24–28 nt), lower annealing Tₘ |
Recommended Primer Sequences for FBXW7
| Target Region | Forward Primer (5′→3′) | Reverse Primer (5′→3′) | Amplicon |
|---|---|---|---|
| Exon 8 (R465) | 5′-GTGTCTATGGTGCCAACTGG-3′ | 5′-GCTCGAGCTGTCTATCCACA-3′ | 173 bp |
| Exon 9 (WD40) | 5′-CCCCATAAGCAGCACATGAG-3′ | 5′-CCTGAGACCCAGTGATTCCT-3′ | 185 bp |
Key SNPs to Avoid in Primer Binding Sites
When designing FBXW7 primers, avoid these clinically significant variants:
- rs121913208 (R465C) — Exon 8 WD40 domain loss-of-function
- rs121913209 (R479Q) — Exon 9 substrate-binding mutation
Clinical Validation Required
All FBXW7 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 FBXW7 Primers Now
Pre-configured with FBXW7-specific parameters. Use our primer design, Tm calculator, and GC content tools for optimal results.