MOLECULAR CHARACTERIZATION AND PCR DIAGNOSIS OF THAILAND DELETION OF ALPHA-GLOBIN GENE-CLUSTER

Thailand deletion of alpha-Thalassemia (thal) 1 involves the zeta 2-, phi zeta 1-, alpha 2-, alpha 1-, and 01-globin genes. In Southeast Asi ans and Taiwanese, this mutation is the second most common long-segmen t deletion of two alpha-globin genes, after the Southeast Asian deleti on, To define the Thailand deletion breakpoints, we used polymerase ch ain reaction (PCR) to amplify the normal-sequence DNA fragments across the breakpoints. The amplified products were sequenced directly or af ter cloning into pGem(R)-3Z or pCR(R)2.1 vectors, Com parison of the n ormal and mutant sequences revealed that the 5' breakpoint lies betwee n nucleotides 1,269 and 1,290 upstream of the initiator codon adenine of the zeta 2-globin gene, and the 3' breakpoint lies between nucleoti des 29,387 and 29,408 downstream of it. A total of 30,677 nucleotides were deleted, Both breakpoints mentioned above lie within the Alu repe titive sequences and an extensive sequence homology is present around the two breakpoints, These findings suggest that homologous recombinat ion is the mechanism by which the deletion occurs, Based on our data, we used three oligonucleotide primers to amplify the regions across th e deletion and its corresponding normal sequence, The feasibility of P CR diagnosis was confirmed in 20 carriers with this deletion. (C) 1998 Wiley-Liss, Inc.