Information on single-nucleotide polymorphisms (SNPs) in hexaploid bread wheat is still scarce. The goal of this study was to detect SNPs in wheat and examine their frequency. Twenty-six bread wheat lines from different origins worldwide were used. Specific PCR-products were obtained from 21 genes and directly sequenced. SNPs were discovered from the alignment of these sequences. The overall sequence polymorphism observed in this sample appears to be low; 64 single-base polymorphisms were detected in approximately 21.5 kb (i.e., 1 SNP every 335 bp). The level of polymorphism is highly variable among the different genes studied. Fifty percent of the genes studied contained no sequence polymorphism, whereas most SNPs detected were located in only 2 genes. As expected, taking into account a synthetic line created with a wild Triticum tauschii parent increases the level of polymorphism (101 SNPs; 1 SNP every 212 bp). The detected SNPs are available at http://urgi.versailles.inra.fr/GnpSNP">http://urgi.versailles.inra.fr/GnpSNP. Data on linkage disequilibrium (LD) are still preliminary. They showed a significant level of LD in the 2 most polymorphic genes. To conclude, the genome size of hexaploid wheat and its low level of polymorphism complicate SNP discovery in this species.

Bioinformatic analyses of maize EST sequences have highlighted large numbers of candidate genes putatively involved in agriculturally important traits. To contribute to ongoing efforts toward mapping of these genes, we used two populations of intermated recombinant inbred lines (IRILs), which allow a higher map resolution than nonintermated RILs. The first panel (IBM), derived from B73 × Mo17, is publicly available from the Maize Genetics Cooperation Stock Center. The second panel (LHRF) was developed from F2 × F252 to map loci monomorphic on IBM. We built framework maps of 237 loci from the IBM panel and 271 loci from the LHRF panel. Both maps were used to place 1454 loci (1056 on map IBM_Gnp2004 and 398 on map LHRF_Gnp2004) that corresponded to 954 cDNA probes previously unmapped. RFLP was mostly used, but PCR-based methods were also performed for some cDNAs to map SNPs. Unlike in usual IRIL-based maps published so far, corrected meiotic centimorgan distances were calculated, taking into account the number of intermating generations undergone by the IRILs. The corrected sizes of our framework maps were 1825 cM for IBM_Gnp2004 and 1862 cM for LHRF_Gnp2004. All loci mapped on LHRF_Gnp2004 were also projected on a consensus map IBMconsensus_Gnp2004. cDNA loci formed clusters near the centromeres except for chromosomes 1 and 8.

Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a group of genetically heterogeneous neurodegenerative disorders characterized by pro- gressive spasticity of the lower limbs. Five AD-HSP loci have been mapped to chromosomes 14q, 2p, 15q, 8q and 12q. The SPG4 locus at 2p21-p22 has been shown to account for approximately 40% of all AD-HSP families. SPG4 encoding spastin, a putative nuclear AAA protein, has recently been identified. Here, sequence analysis of the 17 exons of SPG4 in 87 unrelated AD-HSP patients has resulted in the detection of 34 novel mutations. These SPG4 mutations are scattered along the coding region of the gene and include all types of DNA modification including missense (28%), nonsense (15%) and splice site point (26.5%) mutations as well as deletions (23%) and insertions (7.5%). The clinical analysis of the 238 mutation carriers revealed a high proportion of both asymptomatic carriers (14/238) and patients unaware of symptoms (45/238), and permitted the redefinition of this frequent form of AD-HSP.

A large set of genes was identified in the phytopathogenic fungus Botrytis cinerea by using an expressed sequence tag approach. The fungus was grown in axenic culture and a cDNA library was produced. From this library, 6559 ESTs were obtained. The combined sequences represent 3026 unisequences that corresponds to approximately one-quarter of the estimated total number of genes in B. cinerea. Approximately 18% of the ESTs showed significant similarities with genes coding for proteins with known functions,~56% were similar to genes coding for proteins with unknown functions and ~26% were orphans. A substantial B. cinerea gene inventory including putative virulence factors was therefore obtained and is now available at the Génoplante-Info Database interface (http://urgi.infobiogen.fr///Projects/GPiDB/Interface/).

Motivation: For plant studies, comparative genomics is applied to transfer structural and functional knowledge from model genomes to related genomes.Results: To support the biologist in this comparative genomic task, we have set up an environment for genetic/physical map data management, visualization and comparison: GpiMap.

Génoplante is a partnership program between public French institutes (INRA, CIRAD, IRD and CNRS) and private companies (Biogemma, Bayer CropScience and Bioplante) that aims at developing genome analysis programs for crop species (corn, wheat, rapeseed, sunflower and pea) and model plants ( Arabidopsis and rice). The outputs of these programs form a wealth of information (genomic sequence, transcriptome, proteome, allelic variability, mapping and synteny, and mutation data) and tools (databases, interfaces, analysis software), that are being integrated and made public at the public bioinformatics resource centre of Génoplante: GénoPlante-Info (GPI). This continuous flood of data and tools is regularly updated and will grow continuously during the coming two years. Access to the GPI databases and tools is available at http://genoplante-info.infobiogen.fr/ .

Rickettsia conorii is an obligate intracellular bacterium that causes Mediterranean spotted fever in humans. We determined the 1,268,755-nucleotide complete genome sequence of R. conorii , containing 1374 open reading frames. This genome exhibits 804 of the 834 genes of the previously determined R. prowazekii genome plus 552 supplementary open reading frames and a 10-fold increase in the number of repetitive elements. Despite these differences, the two genomes exhibit a nearly perfect colinearity that allowed the clear identification of different stages of gene alterations with gene remnants and 37 genes split in 105 fragments, of which 59 are transcribed. A 38-kilobase sequence inversion was dated shortly after the divergence of the genus.

Schwartz-Jampel syndrome (SJS1) is a rare autosomal recessive disorder characterized by permanent myotonia (prolonged failure of muscle relaxation) and skeletal dysplasia, resulting in reduced stature, kyphoscoliosis, bowing of the diaphyses and irregular epiphyses1. Electromyographic investigations reveal repetitive muscle discharges, which may originate from both neurogenic and myogenic alterations2,3. We previously localized the SJS1 locus to chromosome 1p34–p36.1 and found no evidence of genetic heterogeneity4,5. Here we describe mutations, including missense and splicing mutations, of the gene encoding perlecan (HSPG2) in three SJS1 families. In so doing, we have identified the first human mutations in HSPG2, which underscore the importance of perlecan not only in maintaining cartilage integrity but also in regulating muscle excitability.

Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a genetically heterogeneous disorder characterized by progressive spasticity of the lower limbs. A major locus (SPG4) causing AD-HSP in about 40% of the families was mapped to chromosome 2p. The analysis of six SPG4-linked AD-HSP families using the RED procedure previously showed the expansion of a CAG repeat in affected individuals. To identify the gene responsible for this form of HSP, we have constructed a 3.5-Mb YAC contig flanked by loci D2S400 and D2S367, have subcloned five of these YACs spanning the candidate region into cosmids, and screened these cosmid libraries for the presence of CAG repeat sequences. Four CAG repeats have been identified but none of them is expanded in 26 patients from 13 SPG4-linked AD-HSP families. A gene map comprising 21 transcripts was established using expressed sequence tags (ESTs) assigned previously to this region of 2p21–p22 with radiation hybrid panels GeneBridge 4 and G3. Full-length cDNAs corresponding to the 14 ESTs mapping to the SPG4 interval flanked by loci D2S352 and D2S2347 were isolated and sequenced. None contains a CAG repeat in its coding sequence. Finally, we have assembled a BAC contig composed of 37 clones that were also screened for the presence of CAG repeats; this failed to detect additional repeats to those identified on YACs.

Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a genetically heterogeneous neurodegenerative disorder characterized by progressive spasticity of the lower limbs. Among the four loci causing AD-HSP identified so far, the SPG4 locus at chromosome 2p21–p22 has been shown to account for 40–50% of all AD-HSP families. Using a positional cloning strategy based on obtaining sequence of the entire SPG4 interval, we identified a candidate gene encoding a new member of the AAA protein family, which we named spastin. Sequence analysis of this gene in seven SPG4 -linked pedigrees revealed several DNA modifications, including missense, nonsense and splice-site mutations. Both SPG4 and its mouse orthologue were shown to be expressed early and ubiquitously in fetal and adult tissues. The sequence homologies and putative subcellular localization of spastin suggest that this ATPase is involved in the assembly or function of nuclear protein complexes.

Familial Mediterranean fever (FMF) is a recessively inherited disorder characterized by attacks of fever and serositis, which affects primarily non-Ashkenazi Jews, Armenians, Turks, and Arabs. We present here a transcriptional map covering the FMF locus that we constructed in the course of the positional cloning of the gene responsible for this disease. This map was established from a contig constructed with YAC, BAC, and cosmid clones and covers about 500 kb of 16p13.3. It contains nine transcriptional units corresponding to known genes or to genes belonging to known gene families, 23 gene fragments characterized by partial sequences, and an endogenous retrovirus sequence. It thus considerably increases the number of genes in this interval and improves our knowledge concerning some of the genes or gene families present in this region. Data accumulated in this region were also used in a comparative study of different methods of exon detection.

Autosomal dominant familial spastic paraplegia (AD-FSP) is a genetically heterogeneous neurodegenerative disorder characterized by progressive spasticity of the lower limbs. Three loci on chromosome 14q (SPG3), 2p (SPG4), and 15q (SPG6) were shown to be responsible for AD-FSP. Analysis of recombination events in three SPG3-linked families allowed us to narrow the critical interval from 9 to 5 cM. An approximately 5-Mb YAC contig comprising 32 clones and 90 STSs was built from D14S301 to D14S991, encompassing this region of 14q21. Fifty-six ESTs assigned previously to this region with radiation hybrid (RH) panels Genebridge 4 and G3 were precisely localized on the YAC contig. The 90 STSs positioned on the contig were tested on the TNG RH panel to compare our YAC-based map with an RH map at a high level of resolution. Comparison between our map and the whole genome mapping data on this interval of chromosome 14q is discussed.

A candidate gene for Branchio-Oto-Renal (BOR) syndrome was identified at chromosome 8q13.3 by positional cloning and shown to underlie the disease. This gene is a human homologue of the Drosophila eyes absent gene (eya), and was therefore called EYA1. A highly conserved 271-amino acid C-terminal region was also found in the products of two other human genes (EYA2 and EYA3), demonstrating the existence of a novel gene family. The expression pattern of the murine EYA1 orthologue, Eya1, suggests a role in the development of all components of the inner ear, from the emergence of the otic placode. In the developing kidney, the expression pattern is indicative of a role for Eya1 in the metanephric cells surrounding the ‘just-divided’ ureteric branches.

Branchio-oto-renal (BOR) syndrome is an autosomal dominant disorder, characterised by the association of branchial, otic and renal anomalies with variable degrees of severity, We have recently identified EYA1, a human homologue of the Drosophila eyes absent gene, as the gene underlying this syndrome, The products of both genes share a highly conserved 271 amino acid C-terminal region (eyaHR). The eyaHR was also found in the products of two other human genes (EYA2 and EYA3), demonstrating the existence of a novel gene family, We report here on the complete genomic structure of EYA1, This gene consists of 16 coding exons and extends over 156 kb, It encodes various alternatively spliced transcripts differing only in their 5' regions, Sequence analysis of the entire EYA1 coding region was performed for 20 unrelated patients affected by BOR syndrome, and six novel mutations were identified, Among these mutations, two are missense mutations, highlighting amino acid residues essential for the function of the EYA1 protein, and one mutation comprises a de novo Alu insertion into an exon, This insertion presumably occurs by retrotransposition, and the mobile Alu element has a poly(A) tail that is unstable throughout generations, To date, 14 mutations have been detected in BOR patients, all of which are different, However, all the mutations are located within or in the immediate vicinity of the eyaHR; the significance of this clustering is discussed.

Familial Mediterranean fever (FMF) is an autosomal recessive disorder characterized by attacks of fever and serositis. In this paper, we define a minimal co-segregating region of 60 kb containing the FMF gene ( MEFV ) and identify four different transcript units within this region. One of these transcripts encodes a new protein (marenostrin) related to the ret-finger protein and to butyrophilin. Four conservative missense variations co-segregating with FMF have been found within the MEFV candidate gene in 85% of the carrier chromosomes. These variations, which cluster at the carboxy terminal domain of the protein, were not present in 308 control chromosomes, including 162 validated non-carriers. We therefore propose that the sequence alterations in the marenostrin protein are responsible for the FMF disease.