tailieunhanh - Novel technique for rapid detection of a-globin gene mutations and deletions

Populations in Southeast AsiaandSouthChinahave highfrequencies ofa-thalassemia caused by a-globin gene mutations and/or deletions. This study was designed to find an efficient and simple diagnostic test for the mutations and deletions. A duplex polymerase chain reaction (PCR)/denaturing high-pressure liquid chromatography (DHPLC) was used to detect the mutations and deletions. A blinded study of 110 samples, which included 92 a-thalassemia samples with various genotypes and 18 normal DNA samples, was carried out by the methods. The duplex PCR products of the sample with known Constand spring mutation (CS)/aa, Quonsze mutation (QS)/ aa, and Weastmead mutation (WS)/aa DNA showed significantly different profiles, which suggests that DHPLC analysis at C can detect potential. | Novel technique for rapid detection of a-globin gene mutations and deletions JINGZHONG LIU XINGYUAN JIA NING TANG XU ZHANG XIAOYI WU REN CAI LIRONG WANG QUANZHANG LIU BAI XIAO JIM ZHU and QINGTAO WANG BEIJING AND GUANGXI CHINA Populations in Southeast Asia and South China have high frequencies of a-thalassemia caused by a-globin gene mutations and or deletions. This study was designed to find an efficient and simple diagnostic test for the mutations and deletions. A duplex polymerase chain reaction PCR denaturing high-pressure liquid chromatography DHPLC was used to detect the mutations and deletions. A blinded study of 110 samples which included 92 a-thalassemia samples with various genotypes and 18 normal DNA samples was carried out by the methods. The duplex PCR products of the sample with known Constand spring mutation CS aa Quonsze mutation QS aa and Weastmead mutation WS aa DNA showed significantly different profiles which suggests that DHPLC analysis at C can detect potential mutations directly. The DHPLC at 50 C analysis can distinguish the --SEA and nondeletional alleles. The new assay is 100 concordant with the original genotype. In conclusion the technique including the duplex PCR assay followed by DHPLC analysis can be used to diagnose a-thalassemia this methodology is simple rapid accurate semiautomatic and high output and thus it is suitable for large-scale screening. Translational Research 2010 155 148-155 Abbreviations CS Constand spring mutation DC dissociation curve analysis DHPLC denaturing high-performance liquid chromatography Duplex PCR duplex polymerase chain reaction GC guanine-cytosine QS Quonsze mutation RDB reverse dot-blot TEAA triethylammonium acetate WS Weastmead mutation a-Thalassemia is the most common recessively inherited hemoglobin Unlike a-thalassemia in which nondeletional mutations predominate most recognized a-thalassemia involve deletion of 1 or both a-globin genes. The a-globin gene cluster is located on .