𧬠Search by Gene Name & Organism
Gene Name / Symbol / Accession Number
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TP53
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Searching NCBI Gene / RefSeq mRNA...
π ORF Search Results & qPCR Design
π§ͺ Verify qPCR primer candidates with Primer-BLAST
Also check GeneQuick β
Open Primer-BLAST pre-configured with product size, Tm, GC%, Intron inclusion, and more using the matched RefSeq mRNA.
π§ͺ Recommended Primer-BLAST Settings
π Primer Parameters
| PCR product size Min / Max | 70 / 200 |
| # of primers to return | 10 |
| Primer Tm Min / Opt / Max | 59.0 / 60.0 / 61.0 Β°C |
| Max Tm difference | 1 |
𧬠Exon / Intron Selection
| Intron inclusion | β Check ON |
π Primer Pair Specificity Checking
| Database | RefSeq mRNA |
| Organism | 9606 (Homo sapiens) |
| Primer specificity stringency | 2 mismatches |
βοΈ Advanced Parameters
| Primer Size Min / Opt / Max | 15 / 20 / 25 |
| Primer GC content Min / Max | 40 % / 60 % |
| Max Poly-X | 2 |
β¦ Relaxation Settings When No Candidates Are Found
If no primers meeting the criteria are found, relax settings one step at a time from the top.
- Max Poly-X: 2 β 3
- 3β²-terminal T: Allow (common in published primers)
- Tm: Widen to Min 58Β°C, Max 62Β°C
- Product size Max: 250 bp (not strongly recommended)
- For sequences without introns: Uncheck Intron inclusion
* Primer performance is only truly known after testing. It's important to set slightly relaxed conditions and select the best candidate from the results.
The above settings are automatically passed to Primer-BLAST when you click "Open Primer-BLAST with this mRNA".