RESUMEN
PURPOSE: Posterior polymorphous corneal dystrophy (PPCD) is an autosomal dominant disorder, affecting both the corneal endothelium and Descemet's membrane. In the Czech Republic, PPCD is one of the most prevalent corneal dystrophies. The purpose of this study was to determine the chromosomal locus of PPCD in two large Czech families, by using linkage analysis. METHODS: Linkage analysis was performed on 52 members of two Czech families with PPCD and polymorphic microsatellite markers and lod scores were calculated. The candidate gene VSX1 was also screened for mutations. RESULTS: Significant lod scores were obtained with microsatellite markers on chromosome 20. Linkage analysis delineated the Czech PPCD locus to a 2.7-cM locus on chromosome 20, region p11.2, between flanking markers D20S48 and D20S139, which excluded VSX1 as the disease-causing gene in both families. In addition, the exclusion of VSX1 was confirmed by sequence analysis. CONCLUSIONS: This study reports the localization of PPCD in patients of Czech origin to chromosome 20 at p11.2. Linkage data and sequence analysis exclude VSX1 as causative of PPCD in two Czech families. This refined locus for PPCD overlaps the congenital hereditary endothelial dystrophy (CHED1) disease interval, and it is possible that these corneal dystrophies are allelic.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos Par 20/genética , Distrofias Hereditarias de la Córnea/genética , Lámina Limitante Posterior/patología , Endotelio Corneal/patología , Proteínas del Ojo/genética , Proteínas de Homeodominio/genética , Distrofias Hereditarias de la Córnea/epidemiología , Distrofias Hereditarias de la Córnea/patología , República Checa/epidemiología , Femenino , Marcadores Genéticos , Humanos , Escala de Lod , Masculino , Linaje , Sitios de Carácter CuantitativoRESUMEN
BACKGROUND: Despite the continuous production of genome sequence for a number of organisms, reliable, comprehensive, and cost effective gene prediction remains problematic. This is particularly true for genomes for which there is not a large collection of known gene sequences, such as the recently published chicken genome. We used the chicken sequence to test comparative and homology-based gene-finding methods followed by experimental validation as an effective genome annotation method. RESULTS: We performed experimental evaluation by RT-PCR of three different computational gene finders, Ensembl, SGP2 and TWINSCAN, applied to the chicken genome. A Venn diagram was computed and each component of it was evaluated. The results showed that de novo comparative methods can identify up to about 700 chicken genes with no previous evidence of expression, and can correctly extend about 40% of homology-based predictions at the 5' end. CONCLUSIONS: De novo comparative gene prediction followed by experimental verification is effective at enhancing the annotation of the newly sequenced genomes provided by standard homology-based methods.
Asunto(s)
Biología Computacional/métodos , Genoma , Animales , Pollos , Mapeo Cromosómico , Cartilla de ADN , ADN Complementario/metabolismo , Interpretación Estadística de Datos , Bases de Datos Genéticas , Exones , Etiquetas de Secuencia Expresada , Perfilación de la Expresión Génica , Intrones , Modelos Estadísticos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de Secuencia , Análisis de Secuencia de ADN , Programas Informáticos , Distribución TisularRESUMEN
We have developed a systematic approach to generating cDNA clones containing full-length open reading frames (ORFs), exploiting knowledge of gene structure from genomic sequence. Each ORF was amplified by PCR from a pool of primary cDNAs, cloned and confirmed by sequencing. We obtained clones representing 70% of genes on human chromosome 22, whereas searching available cDNA clone collections found at best 48% from a single collection and 60% for all collections combined.
Asunto(s)
Clonación Molecular/métodos , Genoma Humano , Genómica/métodos , Sistemas de Lectura Abierta/genética , Proteoma/genética , Cromosomas Humanos Par 22/genética , Biología Computacional , ADN Complementario/genética , Bases de Datos Genéticas , Humanos , Reacción en Cadena de la Polimerasa , Polimorfismo de Nucleótido Simple/genética , Proyectos de Investigación , Análisis de Secuencia de ADNRESUMEN
We report a second-generation gene annotation of human chromosome 22. Using expressed sequence databases, comparative sequence analysis, and experimental verification, we have extended genes, fused previously fragmented structures, and identified new genes. The total length in exons of annotation was increased by 74% over our previously published annotation and includes 546 protein-coding genes and 234 pseudogenes. Thirty-two potential protein-coding annotations are partial copies of other genes, and may represent duplications on an evolutionary path to change or loss of function. We also identified 31 non-protein-coding transcripts, including 16 possible antisense RNAs. By extrapolation, we estimate the human genome contains 29,000-36,000 protein-coding genes, 21,300 pseudogenes, and 1500 antisense RNAs. We suggest that our revised annotation criteria provide a paradigm for future annotation of the human genome.