A physical map of the chromosome 12 centromere.

While current sequencing efforts consider the detection of alpha satellite repeats as logical end points for map construction, detailed maps of most pericentromeric regions are lacking to confirm this hypothesis. Here we identify the different alpha satellite families present at the pericentromeric region of chromosome 12. The order, size and location of these repeats is established using radiation hybrid analysis, pulsed field gel analysis and FISH and the maps are integrated with current sequence information. For the different classes of alpha satellites present at the chromosome 12 centromere the paralogs in the human genome were mapped by FISH. Unique sequences flanking the alpha satellite repeats were identified, some of which are not represented in the current draft sequence. This mapping effort localises the different alpha satellite repeats within the pericentromeric region and anchors them in the current maps. The novel sequences identified may serve as the end point for the ongoing sequencing efforts.

No evidence for the involvement of CAG/CTG repeats from within 18q21.33-q23 in bipolar disorder.

We previously identified 18q21.33-q23 as a candidate region in one BP family and constructed a yeast artificial chromosome (YAC) contig map. Here, we mapped eight known CAG/CTG repeats relative to 18q21.33-q23. We also isolated four CAG/CTG repeats from within the region using CAG/CTG YAC fragmentation, one of which is located in the 5' untranslated region of the CAP2 gene coding for a brain-expressed serine proteinase inhibitor. The triplet repeats located in the 18q21.33-q23 BP candidate region showed no expanded alleles in the linked BP family nor in a BP case-control sample. Moreover, only the CAP2 triplet repeat was polymorphic but no genetic association with BP disorder was observed.

Molecular interpretation of expanded RED products in bipolar disorder by CAG/CTG repeats located at chromosomes 17q and 18q.

Previously we provided evidence that the anticipation observed in bipolar (BP) disorder may be explained by expanded CAG/CTG triplet repeats. Data were generated with the repeat expansion detection (RED) method in a BP case-control sample showing a significant association of BP disorder with expanded CAG/CTG repeats (RED products of 120 bp). In this study we demonstrated that 86% of the RED expansions could be accounted for by the ERDA1 and CTG18.1 CAG/CTG repeats located respectively on chromosomes 17 and 18. Further, significantly different allele distributions were observed for ERDA1, with a larger proportion of BP patients (34.7%) carrying one or two expanded ERDA1 alleles (CAG/CTG repeats >40) than controls (19.2%) (P = 0.032). Also, a negative correlation was observed for ERDA1 between CAG/CTG length and age at onset in affected offspring of eight BP families. Although interesting, these data should be interpreted with caution since the ERDA1 association did not remain significant after correcting for multiple testing. Also, no linkage was observed between BP disorder and expanded ERDA1 alleles in the families.

Molecular genetic analysis of autosomal dominant cerebellar ataxia with retinal degeneration (ADCA type II) caused by CAG triplet repeat expansion.

Autosomal dominant cerebellar ataxia with retinal degeneration (ADCAII) was previously mapped by linkage analysis studies to chromosome 3p12-p21.1 (SCA7). Positional cloning efforts have recently identified a novel gene, SCA7 , containing a translated CAG repeat, expanded in SCA7 patients. We cloned the SCA7 gene from a yeast artificial chromosome (YAC) clone contig spanning the SCA7 candidate region. Using a combination of genomic sequencing and cosmid-based exon trapping, two expressed sequence tags were identified. Sequencing of the corresponding cDNA clones and RT-PCR analysis identified the full-length SCA7 cDNA. Together, our sequence data defined the intron/exon boundaries of the first two coding exons of the SCA7 gene, with the first exon containing the expanded CAG repeat. Further, sequence comparison with the published SCA7 cDNA identified one additional putative exon in the 5'-UTR region of the SCA7 gene. The SCA7 gene was mapped on the YAC contig in the 2.5 cM interval between D3S1600 and D3S1287. In one extended Belgian SCA7 pedigree the expanded alleles ranged from 38 to at least 55 repeats with allele lengths being inversely correlated with onset age of ADCAII symptoms. The SCA7 repeats increased in length in successive generations. Normal alleles had from four to 18 repeats, with 10 repeats being the most common allele.

A radiation hybrid map with 60 loci covering the entire short arm of chromosome 12.

We present a high-resolution radiation hybrid map of the short arm of human chromosome 12 containing 60 loci, including 44 STSs within or closely associated with expressed sequences, 11 highly polymorphic markers, 2 anonymous sequences, 2 subtelomeric sequences, and 1 centromeric sequence. The 60 loci fell into 48 unique retention patterns, providing a comprehensive map covering the entire short arm of chromosome 12 with an average resolution of approximately 800 kb. Twenty-two unique positions were ordered in a 1000:1 framework map with an average resolution of 1.8 Mb. The proposed order is in good agreement with recently published genetic maps, high-resolution FISH maps, and YAC contigs. The noted inconsistencies involved neighboring loci permutations. Our observations further suggest the existence of chromosomal "hot spots" for breakage during irradiation. In three regions an usually high number of breaks was noted between neighboring loci compared to the physical distance derived from existing YAC contigs. Some of these hot spots seem to coincide with known chromosomal aberrations, of which at least two have been involved in the etiology of cancer.

Isolation and regional assignment of human chromosome 12p cDNAs.

We have characterized 117 cDNAs isolated by direct cDNA selection using pools of human chromosome 12p cosmids. Sequencing revealed that 41 clones did overlap with other cDNAs. Of the remaining 76 cDNA sequences, 11 matched previously identified human chromosome 12p genes and 3 matched previously determined cDNA sequences, including the retinoblastoma binding protein 2 (RBBP2), the cyclin-dependent kinase inhibitor KIP1, and an expressed sequence tag. For each of the 76 cDNAs specific selection by a genomic cosmid clone was confirmed. STSs were developed for all cosmids, among them 3 polymorphic simple sequence repeats associated with, respectively, the TNFR related protein, CD27, and SCNN1. Regional assignment of the STSs by PCR analysis with somatic cell hybrids and fluorescence in situ hybridization showed that the majority of the loci map to chromosome 12p13, similar to the distribution of the known 12p genes. Evidence was found for the duplication on 12p of a region containing a polymorphic simple sequence repeat and sequences of two different cDNAs.

Correlations between triplet repeat expansion and clinical features in Huntington's disease.

OBJECTIVE: To investigate possible correlations between the length of the (CAG)n trinucleotide repeat in Hungtington's disease gene IT15 and clinical features (age at onset, symptoms at onset, and mode of progression) in Huntington's disease. DESIGN: In 59 patients with Huntington's disease, the expansion of the (CAG)n trinucleotide repeat was determined and clinical data were obtained retrospectively. SETTING: The Center for Human Genetics, affiliated with a university hospital. PATIENTS: All patients belonged to an initial group of 248 individuals tested in an indirect predictive testing procedure. RESULTS: A good correlation was found between the expansion of the (CAG)n trinucleotide repeat and the age at onset (r = -.71). No correlation was found between the repeat length of the normal allele and the age at onset. No correlations were found between repeat expansion and other clinical features, such as the nature of the symptoms at onset (neurologic, psychiatric/cognitive, or both) and the mode of progression. CONCLUSION: Factors that determine the nature of symptoms at onset and the mode of progression of Huntington's disease seem to be operating independently of the (CAG)n trinucleotide repeat in gene IT15.