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Table 2 Genotypes of samples used in the development and validation of the alpha globin gene ratio analysis copy enumeration PCR assay (GRACE-PCR)

From: Development and validation of a high throughput, closed tube method for the determination of haemoglobin alpha gene (HBA1 and HBA2) numbers by gene ratio assay copy enumeration-PCR (GRACE-PCR)

α-Globin StripAssay GRACE-PCR Screening Test
Genotype Strip Assay (n) PCR 1 (n) PCR 2 (n) CLCN7:HBA1 Ratio HBA1:HBA2 Ratio HBA1 Copies HBA2 Copies
1. αα/αα (wild type) 54 30 24 1:1 1:1 2 2
2. αα/-α3.7 48 29 19 1:1 2:1 2 (a) 1
3. -α3.7/-α3.7 36 20 16 1:1 1:0 2 (a) 0
4. αα/-α4.2 2 2 0 1:1 2:1 2 1
5. -α4.2/-α4.2 1 1 0 1:1 1:0 2 0
6. -α3.7/-α4.2 3 3 0 1:1 1:0 2 (a) 0
7. -α3.7/--SEA 6 3 3 2:1 1:0 1 (a) 0
8. αα/--Med 2 2 0 2:1 1:1 1 1
9. αα/--Fil 2 2 0 2:1 1:1 1 1
10. -(α)20.5 1 1 0 1:1 2:1 2 (b) 1
11. αα/αααanti3.7 2 2 0 1:1 2:3 2 3
12. -α3.7Icariaα (c) 1 1 0 1:1 2:1 2 (a) 1
13. αα/αpoly-A1α (c) 2 2 0 1:1 1:1 2 2
14. αα/αpoly-A2α (c) 1 1 0 1:1 1:1 2 2
15. -α3.7polyA-1α (c) 4 4 0 1:1 2:1 2 (a) 1
16. αα/αConstant Springα (c) 1 1 0 1:1 1:1 2 2
17. αConstant Springα/αConstant Springα (c) 1 1 0 1:1 1:1 2 2
Total number of samples 167 105 62  
  1. The GRACE-PCR assay was assessed using samples with previously determined (PCR1) and unknown (PCR 2) genotypes. GRACE-PCR was able to detect all 108 samples identified by the commercial α-globin StripAssay as having α-globin gene rearrangements (genotypes 2 to12)
  2. (a) Due to the positioning of primers at the 3′ ends of the α-globin genes, the hybrid -α3.7 gene is counted as an HBA1 gene in this assay
  3. (b) The -(α)20.5 deletion does not extend to the 3′ end on the HBA1 gene targeted by the GRACE-PCR screening test primers, consequently only the deletion of the HBA2 gene is detected
  4. (c) Samples with point mutations (genotypes 12 to 17) were included to ensure that these point mutations did not affect the gene-copy number determination of the GRACE-PCR assay