The primary objective of this investigation was to determine the molecular basis of
ß-thalassemia in two different families so that parental diagnosis could be undertaken and genetic counselling offered from a position of knowledge rather than statistical probability.
The family groups (one Chinese and the other European) consisted of a couple, together with their parents. In both couples each partner had been diagnosed as being a carrier of
ß-thalassemia on haematological grounds, giving rise to a 1:4 probability that the offspring might have severe ß-thalassemia.
After DNA extraction, the entire ß-globin gene was amplified by PCR, and a third primer used for sequence analysis. In Case 1, the Chinese couple, both partners were found to have mutations in the critical TATA box region which is involved in the regulation of transcription. One partner (11-1) had a -29 A→G mutation while the other (11-2) had a -28 A→G mutation. These findings were confirmed using allele-specific probes and dot blot analysis. Dot blot analysis of amplified DNA of the chorionic villus sample (111-1) from patient 11-2 indicated that the fetus was a heterozygote for the -29 A→G.
In Case 2, the European family, one partner (11-3) was found to have a mutation of C→T at IVS 2 nt 654 while the other partner (11-4) had a mutation of CAG→TAG at codon ß39. The former mutation results in an abnormal splicing event, while the latter results in premature chain termination. Again these findings were confirmed by dot blot hybridisation. Dot blot analysis of the amplified DNA of the chorionic villus sample (111-2) from patient 11-4 indicated the fetus was a mixed heterozygote for both these mutations. An incidental finding in both families was the presence of a polymorphism in the histidine codon at ß2 (CAC→CAT).
During this investigation a blood sample was referred for a patient with a polycythemia that was due to the presence of a high oxygen affinity haemoglobin with an unknown ß-chain mutation. DNA sequence analysis of Patient A in this instance revealed a mutation of ß36 Pro (CCT)→Thr (ACT). This proline residue is essential in maintaining the stability of deoxyhaemoglobin and its replacement by threonine destabilises the T form of the molecule favouring the R form or oxyhaemoglobin.[Show full abstract]
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