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Fig. 4. Fragment-size analysis using slab gels and autoradiography: Fragile X. The autoradiograph shows PCR products using primers flanking the CCG repeat 5' of the FMR1 gene. Sizes of patient alleles are determined in reference to the A lanes from a sequencing reaction using phage M13 as the template. The arrows indicate the position of the 204 and 254 base fragments of the sequencing ladder. Lanes 1, 4, and 6 show hemizygous males with amplicons of 200 bases, or 29 repeats. Lanes 2, 5, and 7 are from males with 30 repeats, Lane 3 shows a male with 43 repeats. Lane 8 shows a female heterozygous for a premutation allele of 52 repeats and a normal allele of 31 repeats. Lane 9 shows a control female sample with 30 and 87 repeats. Lane 10 is a no-DNA amplification control. (Courtesy of the Mayo Clinic Molecular Genetics Laboratory.)

5.2. SINGLE-STRANDED CONFORMATIONAL POLYMORPHISM ANALYSIS FOR DETECTION OF MUTATIONS AND SEQUENCE VARIANTS Single-stranded conformational polymorphism (SSCP) analysis is one of the most widely used mutation scanning systems for identification of unknown sequence variations. The reasons for its popularity are its high sensitivity to the presence of sequence variations and its technical simplicity. The extent of the popularity of SSCP can be appreciated from noting that a recent PubMed search of the term "SSCP" gave over 8000 hits. That many investigators worldwide have utilized this technique is appreciated by noting the number of PubMed citations over the years since 1989 (Fig. 6). The technique, developed in Hayashi's laboratory and first reported in 1989, involves PCR amplification of the region

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