Development of a comprehensive molecular diagnosis system for neurological diseases based on DNAmicroarrays.

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This project focused on development of a comprehensive molecular diagnosis system for neurological diseases based on DNA microarrays. lb accomplish this aim, we have developed 1. DNA microarray-based comprehensive resequencing system, and 2. high density array-CGH system to accomplish detection of deletions/multiplications and identification of breakpoints We have developed DNA microarray-based resequencing system for amyotrophic lateral sclerosis, Parkinson disease, adrenoleukodystrophy, and familial spastic paraplegia. With this system, we have shown DNA microarray-based resequencing system is highly efficient to identify point mutations. Although DNA resequencing microarrays are quite effective for identification of point mutations, they are inefficient for detection of deletions or multiplication. To overcome this problem, we have newly developed high density array-CGH system to allow detection of deletions/multiplications of PARK2 gene with the resolution of 100-200bp. Since the resolution is extremely high, the junction segments can be easily amplified by PCR employing PCR primers flanking the deletions/multiplications, allowing identification of breakpoints of deletions/multiplications on nucleotide levels. We applied this system for investigation of the mechanisms of deletions/multiplications of PARK2 in patients with autosomal recessive juvenile Parkinsonism (AR JP). We have determined deletions/multiplications of 299 alleles. The breakpoints clustered in a narrow region of PARK2 that coincides with the center of FRA6E (common fragile site). Indeed analysis of 120 cancer cell lines allowed 31 deletions/multiplications and the distribution is quite similar to that found in ARJP Taken together these studies demonstrate that common mechanisms underlie the of deletions/multiplications in the germline mutations (AR-JP) as well as somatic mutations (cancer cell lines).