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Two novel beta-thalassemia mutations are described. The first mutation, found in an Italian family, is a G----A substitution in nucleotide (nt) +22 relative to the beta-globin gene Cap site. This mutation creates a cryptic ATG initiation codon, the utilization of which for translation would result in premature termination 36 bp 3' downstream. The second mutation, found in an Irish family, is a T----C substitution in nt +1570, or 12 bp 5' upstream of the AATAAA polyadenylation signal in the 3' noncoding region. It is postulated that this mutation leads to destabilization of the encoded beta-globin mRNA.

We report a relatively mild phenotype associated with two siblings who are compound heterozygotes for Hb S and a beta zero-thalassemia mutation due to a approximately 1.4-kb deletion of the 5' region of the beta-globin gene. Each is found to have unusually high levels of Hb A2 and Hb F, accounting for more than 20% of the total hemoglobin. These may interfere with intracellular Hb S polymerization, thus leading to a mild clinical course.

The 1.8-kb beta-globin gene fragments of DNAs from individuals heterozygous for nine different beta-thalassemia mutations involving 1, 2, 3, 4, or 25 basepair (bp) insertions or deletions were amplified by the polymerase chain reaction (PCR). The PCR products were subjected to electrophoresis on aqueous 8% polyacrylamide gel. In each heterozygote with either a 2 to 25 bp deletion, but not with a 1 bp insertion, two slower migrating bands representing heteroduplexes in addition to the 1.8-kb homoduplex band were seen. The electrophoretic positions of these slower migrating bands were characteristic of each mutation studied. By co-amplification with known normal DNA, it was also possible to distinguish DNAs from normal individuals and from individuals who are homozygous for the small insertion/deletion mutations. These studies demonstrate that the heteroduplex formation generated in PCR can be applied as a simple method in the diagnosis of insertion/deletion mutations involving 2 to 25 bp in beta-thalassemias as well as in other genetic disorders.

A new beta zero-thalassemia mutation, a frameshift mutation with an insertion of a single cytosine nucleotide in codon 27-28, is described. The propositus, who is compound heterozygous for this mutation and the IVSII-654 C----T beta zero-thalassemia mutation, has the phenotype of severe beta-thalassemia major.

We identify and characterize a novel beta 0-thalassemia mutation that is associated with an unusually high level of hemoglobin (Hb) A2 in the heterozygote. This newly discovered mutation is caused by a 532-basepair deletion that extends from positions -454 to + 78 relative to the mRNA cap site of the beta-globin gene. The propositi are 9-month-old fraternal twins. One of the twins is a compound heterozygote for the deletion and Hb S, the other is a compound heterozygote for the deletion and Hb C.

We used denaturing gradient gel electrophoresis to detect the beta-thalassemia mutations in the Chinese population. By amplifying the beta-globin gene in four separate fragments and electrophoresing the amplified DNA in two gels, we were able to distinguish all the 12 known mutations on the basis of the mobility of the homoduplexes and heteroduplexes. We conclude that denaturing gradient gel electrophoresis offers a nonradioactive means of detecting multiple mutations in genetic disorders.

We describe a simple approach for detecting beta-thalassemia mutations in geographic areas such as southern China where multiple mutations are known to occur. Segments of the beta-globin gene were amplified in vitro by using the polymerase chain reaction. Dot blot hybridization of the amplified DNA with oligonucleotide probes corresponding to the six mutations found in southern China could directly identify the mutations causing beta-thalassemia in the affected families. The increased number of target sequences after amplification allows the use of 35S-labeled probes, which are reusable for up to 3 months. The mutations can be determined in two days.

We used in vitro DNA amplification by the polymerase chain reaction and nonradioactive probes for prenatal diagnosis of beta thalassemia in Chinese from the Guangdong province. Exact molecular diagnoses were made in all 20 fetuses studied over a 6-month period. We conclude that this method of prenatal diagnosis for beta thalassemia is a viable approach in many parts of the world where this disease is common.

The phenotype of beta thalassemia can be caused by over 40 different mutations. To set up a prenatal diagnosis program using DNA analysis, it is important to determine the type and frequency of mutation in a particular geographic area. We have delineated the molecular lesions that cause beta thalassemia in the Guangdong province of China, and found six mutations in four different haplotypes. The surprising finding that five of these mutations each occur in two different haplotypes suggests the occurrence of crossing over or gene conversion events at the beta-globin locus. The delineation of the haplotypes and mutations will permit the choice of the appropriate probes for prenatal detection of beta thalassemia in this part of China.

During the course of prenatal diagnosis for beta-thalassemia in Chinese patients, we encountered a mutation that was not detectable by oligonucleotides for the known Chinese mutations. Amplification of the beta-globin gene by the polymerase chain reaction and direct DNA sequencing revealed a previously undescribed -30 TATA box mutation which was carried by the father. Prenatal diagnosis was achieved, and the fetus did not inherit this beta-thalassemia allele.

A total of 72 chromosomes from 36 Indonesian patients, 23 with beta-thalassemia major and 13 with Hb E-beta-thalassemia, were analyzed by specific oligonucleotide hybridization after DNA amplification. Thirteen had the beta E mutation (codon 26 GAG----AAG). Of the 59-beta-thalassemic chromosomes, 32 were of the variant IVS-1 nt5 (G----C). Seven had the mutation IVS-2 nt654 (C----T), one had the mutation codon 41/42 (deletion CTTT), and one had the mutation codon 17 (AAG----TAG). Another six with the mutation IVS-1 nt1 (G----T), one with the mutation IVS-1 nt1 (G----A), four with the mutation codon 15 (TGG----TAG), one with a mutation codon 30 (AGG----ACG), and one with a mutation codon 35 (deletion C) were first identified by direct sequencing of a patient's genomic DNA followed by further hybridizing other patients' DNA with the appropriate oligonucleotide probes. Five did not carry the common mutations previously described in Asian populations. The four most prevalent mutations encountered made up 83% of the total number of beta-thalassemic chromosomes studied. The most common mutation, IVS-1 nt5 (G----C), was mostly associated with two different haplotypes.