RESUMEN
We describe a technology, the NanoString nCounter gene expression system, which captures and counts individual mRNA transcripts. Advantages over existing platforms include direct measurement of mRNA expression levels without enzymatic reactions or bias, sensitivity coupled with high multiplex capability, and digital readout. Experiments performed on 509 human genes yielded a replicate correlation coefficient of 0.999, a detection limit between 0.1 fM and 0.5 fM, and a linear dynamic range of over 500-fold. Comparison of the NanoString nCounter gene expression system with microarrays and TaqMan PCR demonstrated that the nCounter system is more sensitive than microarrays and similar in sensitivity to real-time PCR. Finally, a comparison of transcript levels for 21 genes across seven samples measured by the nCounter system and SYBR Green real-time PCR demonstrated similar patterns of gene expression at all transcript levels.
Asunto(s)
Sondas de ADN/metabolismo , Perfilación de la Expresión Génica/métodos , Nanotecnología/instrumentación , Nanotecnología/métodos , Línea Celular , Color , Sondas de ADN/genética , Biblioteca de Genes , Genes Reporteros , Humanos , Procesamiento de Imagen Asistido por Computador , Análisis de Secuencia por Matrices de Oligonucleótidos , Poliovirus , Reacción en Cadena de la Polimerasa , ARN Mensajero/análisis , ARN Mensajero/genética , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
Chromosome 14 is one of five acrocentric chromosomes in the human genome. These chromosomes are characterized by a heterochromatic short arm that contains essentially ribosomal RNA genes, and a euchromatic long arm in which most, if not all, of the protein-coding genes are located. The finished sequence of human chromosome 14 comprises 87,410,661 base pairs, representing 100% of its euchromatic portion, in a single continuous segment covering the entire long arm with no gaps. Two loci of crucial importance for the immune system, as well as more than 60 disease genes, have been localized so far on chromosome 14. We identified 1,050 genes and gene fragments, and 393 pseudogenes. On the basis of comparisons with other vertebrate genomes, we estimate that more than 96% of the chromosome 14 genes have been annotated. From an analysis of the CpG island occurrences, we estimate that 70% of these annotated genes are complete at their 5' end.