In a first step, PCR amplification is performed across the CCTG repeat.[9] For genomic Southern analysis, DNA is digested with BsoBl or EcoR1.[2] Because of the size and somatic instability of the CCTG repeat, Southern analysis fails to detect expansions in 20% of the known carriers.[2] Expanded alleles appear as single discrete bands, multiple bands, or smears. Contrary to other expansion disorders, ZNF9 expansions are almost always less intense than normal alleles.[2] To improve detection of CCTG repeat expansions, a repeat assay has been developed, amplifying the region containing the CCTG expansion by use of the primers CL3N58-D R (5'-GGCCTTATAACCATG-CAAATG-3'), JJP4CAGG (5'-TACGCATCCGAGTTT-GAGACGCAGGCAGGCAGGCAGGCA-GG-5), and JJP3 (5'-TACGCATCCGAGTTTGAGACG-3') followed by Southern analysis of the PCR products probed with an internal probe. By using a PCR primer that primes from multiple sites within the CCTG repeat, expansions are detected by the presence of a smear of products with a molecular weight higher than in control lanes.[2] To ensure specificity, repeat assay PCR products are transferred to a nylon membrane and probed with an internal oligonucle-otide probe.[2] The repeat assay cannot be used to determine the CCTG repeat length.

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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