Primer sequences for α-thalassaemia multiplex gap-PCR and multiplex amplification refractory mutation system (ARMS) assay
| Allele detected | Name | Primer sequences (5′ → 3′) | Conc (μM) | Fragment (bp) |
|---|---|---|---|---|
| *This fragment migrates at ∼1200 bp on non-denaturing polyacrylamide gels. | ||||
| α-thalassaemia multiplex gap-PCR | ||||
| LIS 3′UTR fragment | LIS1 (forward) | ATACCATGGTTACCCCATTGAGC | 0.5 | 2350 |
| (internal control) | LIS1 (reverse) | AGGGCTCATTACATGTGGACCC | 0.5 | |
| −α3.7 | α2/3.7 (forward) | CCCCTCGCCAAGTCCACCC | 0.2 | 2022/2029 |
| 3.7 (reverse) | AAAGCACTCTAGGGTCCAGCG | 0.2 | ||
| α2 gene | α2/3.7 (forward) | as above | – | 1800 |
| α2 (reverse) | AGACCAGGAAGGGCCGGTG | 0.2 | ||
| −α4.2 | 4.2 (forward) | GGTTTACCCATGTGGTGCCTC | 0.5 | 1628 |
| 4.2 (reverse) | CCCGTTGGATCTTCTCATTTCCC | 0.5 | ||
| −−SEA | SEA (forward) | CGATCTGGGCTCTGTGTTCTC | 0.2 | 1349 |
| SEA (reverse) | AGCCCACGTTGTGTTCATGGC | 0.2 | ||
| α-thalassaemia multiplex ARMS assay | ||||
| ζ2-globin gene fragment | ENG12 (forward) | GGATCCACGCAGTGCTAGAAG | 0.294 | 930* |
| (internal control) | ENG21 (reverse) | GTAGAGATGGTGTTTTGCCATGT | 0.294 | |
| codon 30 (ΔGAG) | ARMSC30 (forward) | GTATGGTGCGGAGGCCCTGAG | 0.074 | 772 |
| BE17 (reverse) | CCATTGTTGGCACATTCCGGGA | 0.140 | ||
| codon 125 (CTG→CCG) | ARMSC125 (forward) | CACCCCTGCGGTGCACGCCTCACC | 0.013 | 234 |
| Hb Quong Sze | BE17 (reverse) | as above | – | |
| Termination codon (TAA→CAA) | ARMSC142 (forward) | CCGTGCTGACCTCCAAATACGGTC | 0.096 | 184 |
| Hb Constant Spring | BE17 (reverse) | as above | – | |