An improved fragile X Southern blot probe without the CGGs eliminates background bands.
The largest of the commonly used probes for Southern blot diagnosis of fragile X mental retardation syndrome spans the CGG repeat cluster in the FMR-1 gene. This probe causes the appearance of 'common' or 'constant' background bands which occasionally complicate the interpretation of autoradiographic results. By removing a 357 bp Sphl to Nhel fragment containing the CGGs from the probe pE5.1, we constructed a probe which eliminates the background bands yet allows the use of a large probe (4.8 kb) to detect changes in the diagnostic 5.2 kb genomic EcoRl band. This CGG-deficient probe has been used in routine diagnostic cases as well as in second round testing of pE5.1-probed cases where enlarged mutant bands are suspected to comigrate with the background bands.
Department of Molecular Biology, Massachusetts General Hospital, Boston 02114.
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Davidow
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Barberio
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Cao
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McCormick
D001345:Autoradiography; D015139:Blotting, Southern / Q000379:methods*; D003597:Cytosine Nucleotides / Q000032:analysis*; D004247:DNA / Q000032:analysis* / Q000235:genetics; D005600:Fragile X Syndrome / Q000235:genetics*; D006150:Guanine Nucleotides / Q000032:analysis*; D006801:Humans; D009154:Mutation; D016133:Polymerase Chain Reaction
D003597:Cytosine Nucleotides; D006150:Guanine Nucleotides; D004247:DNA