RESUMEN
We have characterized 11 overlapping yeast artificial chromosomes (YACs) in the 1p13 region, 8 of them containing the human nerve growth factor (NGF) gene (HGMW-approved symbol NGFB). Sequence-tagged sites (STSs) corresponding to YAC extremities have been designed and used for chromosome assignment on a panel of monochromosomic somatic cell hybrids to check for YAC chimerism, in parallel with analyses by fluorescence in situ hybridization. Determination of end STS content and restriction mapping of the YACs led to the construction of a 3-Mb YAC contig. Four microsatellite markers from the Généthon collection and seven genes known to map to the 1p13 region have been ordered on the contig around the NGF gene. A new gene transcript from the Genexpress catalog has been localized on the contig. This work provides an integrated physical, genetic, and genic map of this chromosome 1 region. It constitutes a basis for determining the structure of the NGF gene and for further characterizing its genic environment.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos Par 1/genética , Factores de Crecimiento Nervioso/genética , Animales , Secuencia de Bases , Quimera/genética , Mapeo Cromosómico/métodos , Cromosomas Artificiales de Levadura , Cricetinae , Cartilla de ADN/genética , Marcadores Genéticos , Humanos , Células Híbridas , Hibridación Fluorescente in Situ , Ratones , Repeticiones de Microsatélite , Datos de Secuencia Molecular , Mapeo Restrictivo , Lugares Marcados de SecuenciaRESUMEN
We have tested 80 expressed sequence-tagged site (eSTS) markers assigned to human chromosome 11 by the Genexpress program on a panel of somatic cell hybrids containing parts of this chromosome, characterized by cytogenetic data, reference markers, and with respect to the Généthon microsatellite genetic map. Sixty-eight new gene transcripts have been assigned to 25 subregions, one of which was newly defined by five of the eSTS markers. The markers are distributed on the short and long arms in agreement with their physical length. The genic map thus obtained has been integrated with the cytogenetic, genetic, and disease maps. Two eSTS markers have been further mapped with respect to a yeast artificial chromosome (YAC) contig close to the brain-derived neurotrophic factor (BDNF) gene and thus provide potential candidate genes for the mental retardation phenotype of WAGR (Wilms' tumor, aniridia, genitourinary abnormalities and mental retardation) syndrome. Altogether, the 68 new gene transcripts localized here represent more than a threefold increase in the number of unknown regionalized genes that could reveal potential candidate genes for the numerous orphan pathologies associated with chromosome 11.
Asunto(s)
Cromosomas Humanos Par 11/genética , Animales , Factor Neurotrófico Derivado del Encéfalo , Cromosomas Artificiales de Levadura/genética , ADN Complementario/genética , Expresión Génica , Biblioteca de Genes , Genes , Enfermedades Genéticas Congénitas/genética , Marcadores Genéticos , Humanos , Células Híbridas , Lactante , Ratones , Datos de Secuencia Molecular , Proteínas Musculares/genética , Proteínas del Tejido Nervioso/genética , ARN Mensajero/genética , Lugares Marcados de SecuenciaRESUMEN
WAGR (Wilms tumor, aniridia, genito-urinary abnormalities, mental retardation) syndrome in humans is associated with deletions of the 11p13 region. The brain-derived neurotrophic factor (BDNF) gene maps to this region, and its deletion seems to contribute to the severity of the patients' mental retardation. Yeast artificial chromosomes (YACs) carrying the BDNF gene have been isolated and characterized. Localization of two known exons of this gene leads to a minimal estimation of its size of about 40 kb. Chimerism of the BDNF YACs has been investigated by fluorescence in situ hybridization and chromosome assignment on somatic cell hybrids. Using the BDNF gene, YAC end sequence tagged sites (STS), and Généthon microsatellite markers, we constructed a 1.7-Mb contig and refined the cytogenetic map at 11p13. The resulting integrated physical, genetic, and cytogenetic map constitutes a resource for the characterization of genes that may be involved in the WAGR syndrome.
Asunto(s)
Cromosomas Artificiales de Levadura , Cromosomas Humanos Par 11 , Factores de Crecimiento Nervioso/genética , Proteínas del Tejido Nervioso/genética , Síndrome WAGR/genética , Secuencia de Bases , Factor Neurotrófico Derivado del Encéfalo , Mapeo Cromosómico , Cartilla de ADN , Humanos , Células Híbridas , Hibridación Fluorescente in Situ , Datos de Secuencia Molecular , Lugares Marcados de SecuenciaRESUMEN
Using a panel of 25 somatic cell hybrids, we have regionally localized 112 microsatellite markers generated by Généthon and assigned to chromosome 11. A genetic map of 74 of them was produced using linkage analysis of the eight largest CEPH (Centre d'Etude du Polymorphisme Humain) families. They could be ordered on chromosome 11 with an average distance of 2.1 cM. The tight correlation observed between the genetic order and the physical assignment of these microsatellites reinforces the genetic map data. These newly localized markers identified by the PCR method using a standardized protocol represent useful tools for mapping YAC clones and establishing YAC contigs and for studying genetic diseases or cancers associated with specific genes and/or germinal/somatic rearrangements of chromosome 11.
Asunto(s)
Cromosomas Humanos Par 11 , Animales , Secuencia de Bases , Síndrome de Beckwith-Wiedemann/genética , Neoplasias Óseas/genética , Mapeo Cromosómico , Cromosomas Artificiales de Levadura , Clonación Molecular/métodos , Cartilla de ADN , Desoxirribonucleasas de Localización Especificada Tipo II , Marcadores Genéticos , Humanos , Células Híbridas , Leucemia/genética , Datos de Secuencia Molecular , Roedores , Sarcoma de Ewing/genética , Esquizofrenia/genética , Programas Informáticos , Síndrome WAGR/genéticaRESUMEN
An NGF cDNA containing the 5' exons of the nerve growth factor (NGF) messenger was obtained from chicken heart mRNA using the anchored polymerase chain reaction technique. Alignment of the chicken with the corresponding murine and human sequences reveals interspecies similarities. A sequence corresponding to an exon found only in the NGF messenger, which is abundant in the submaxillary gland of the male mouse, is present in the chicken NGF cDNA. The first non-coding exons of the NGF gene are much less conserved between chicken and mouse or human than the region of the last exon encoding the mature protein. After the cloning of the chicken NGF gene from a cosmid library, the chicken NGF exons have been located within 20 kb of DNA. The chicken NGF gene is therefore shorter than its murine counterpart which spans more than 43 kb. Furthermore, the organization of the chicken and murine NGF genes markedly differs in their 5' portion.