Detection of a novel double mutation in a beta-thalassemia patient by RNA single-stranded conformation polymorphisms and direct sequencing.

Applications of various molecular methods for the analysis of mutations in the @-globin gene has led to the identification of nearly 100 different @-thalassemia mutations in the world.’ Recently, two methods have been described for mutation detection that rely on the analysis of single-stranded conformation polymorphisms (sscp) ofDNA2 or of RNA.’ We find that DNA s s c p and RNA sscp analysis both appear to be capable Of detecting mutations in the @-globin gene but RNA S s c p analysis a P pears to be more informative. We have been able to identify a @-thalFig 1. DNA sscp analysis of a region of8-globin gene by PCR. N1 to N4 represent four normal volunteers and the rest represent samples heterozygous for the iVS 1 nt 5 (G to C change). sam,,les with the mutation show shift in mobility of the lower band assemia gene with two mutations: one at codon 30 (a G to c change), and another at intervening sequence I (IVS I ) nt 5 (G to C change) in one individual by RNA SSCP analysis. To our knowledge this is the first time a double mutation has been found for the N1 N2 N3 N4 consensus splice donor site in the same patient. compared with the bands seen for the normal samples.