Figure 3.
Genotyping study of the mutation that results in hemoglobin M Iwate. (A) Electrophoregram corresponding to the sequencing of exon 2 of the alpha2-globin gene of the proband, showing the heterozygous mutation CAC>TAC at codon 87 (His>Tyr); (B) Electrophoresis of DNA in 1% agarose gel stained with ethidium bromide showing PCR product after restriction digestion with the enzyme RsaI. The largest fragment with 1803 bp does not contain any restriction site for RsaI and corresponds to the amplification of HBA2 without the M Iwate mutation and with the C-polymorphism at site rs2541669 (NG_000006.1:g.33004C>T); the fragments with 1544 pb and 269 bp result from the rs2541669 T-polymorphism and HBA2 without M Iwate mutation; fragments with 1150 and 394 result from the action of RsaI on the new restriction site created by the M Iwate mutation. Specific amplification of HBA2 was performed with alpha2/3.7-F and alpha2-R primers.3
MM = 250 bp molecular marker; bp = base pairs.