A diagram and RT-PCR results showing aberrant splicing as seen in humans of the human βIVS-2-654 transcript in theHbb^th-4/Hbb⁺ thalassemic mice. (A) The diagram shows the human βIVS-2-654 gene with aberrantly spliced β globin mRNA produced from the mutant gene compared with the correctly spliced β globin mRNA that would have been produced from a wild-type gene. The thick line between nucleotides 580 and 652 shows the region of IVS-2 that is maintained in the βIVS-2-654 mRNA. RT-PCR primer i and iii are shown at the location and in the direction in which they anneal to the RNA. Primer i anneals to sequences within the second exon of human β globin, and primer iii anneals to two bases of the sequence at the end of the second exon and 29 bases of the region of the second intron that is maintained with this mutation in humans. (B) An autoradiograph of polyacrylamide gel electrophoresis of RT-PCR with primers i and iii on RNA from a heterozygous huβ^s/Hbb⁺ mouse (lane 1), a heterozygous Hbb^th-4/Hbb⁺ thalassemic mouse (lane 2), a HeLa cell line transfected with a βIVS-2-654 gene (lane 3), and a normal human (lane 4) is shown. Aberrantly spliced β globin mRNA RT-PCR product is 233-bp, as labeled. Correctly spliced β globin mRNA does not amplify with primers i and iii.