RESUMO
The mammalian central nervous system (CNS) contains a remarkable array of neural cells, each with a complex pattern of connections that together generate perceptions and higher brain functions. Here we describe a large-scale screen to create an atlas of CNS gene expression at the cellular level, and to provide a library of verified bacterial artificial chromosome (BAC) vectors and transgenic mouse lines that offer experimental access to CNS regions, cell classes and pathways. We illustrate the use of this atlas to derive novel insights into gene function in neural cells, and into principal steps of CNS development. The atlas, library of BAC vectors and BAC transgenic mice generated in this screen provide a rich resource that allows a broad array of investigations not previously available to the neuroscience community.
Assuntos
Sistema Nervoso Central/metabolismo , Cromossomos Artificiais Bacterianos/genética , Perfilação da Expressão Gênica , Biblioteca Gênica , Genes Reporter/genética , Transgenes/genética , Animais , Axônios/metabolismo , Diferenciação Celular , Linhagem da Célula , Movimento Celular , Sistema Nervoso Central/citologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 1 , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica no Desenvolvimento , Vetores Genéticos/genética , Proteínas de Fluorescência Verde , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proteínas com Homeodomínio LIM , Proteínas Luminescentes/análise , Proteínas Luminescentes/genética , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurociências/métodos , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/metabolismo , Sinapses/metabolismo , Fatores de TranscriçãoRESUMO
Transgenic mouse production via pronuclear microinjection is a complex process consisting of a number of sequential steps. Many different factors contribute to the effectiveness of each step and thus influence the overall efficiency of transgenic mouse production. The response of egg donor females to superovulation, the fertilization rate, egg survival after injection, ability of manipulated embryos to implant and develop to term, and concentration and purity of the injected DNA all contribute to transgenic production efficiency. We evaluated and compared the efficiency of transgenic mouse production using four different egg donor mouse strains: B6D2/F1 hybrids, Swiss Webster (SW) outbred, and inbred FVB/N and C57BL/6. The data included experiments involving approximately 350 DNA transgene constructs performed by a high capacity core transgenic mouse facility. Significant influences of particular genetic backgrounds on the efficiency of different steps of the production process were found. Except for egg production, FVB/N mice consistently produced the highest efficiency of transgenic mouse production at each step of the process. B6D2/F2 hybrid eggs are also quite efficient, but lyze more frequently than FVB/N eggs after DNA microinjection. SW eggs on the other hand block at the 1-cell stage more often than eggs from the other strains. Finally, using C57BL/6 eggs the main limiting factor is that the fetuses derived from injected eggs do not develop to term as often as the other strains. Based on our studies, the procedure for transgenic mouse production can be modified for each egg donor strain in order to overcome any deficiencies, and thus to increase the overall efficiency of transgenic mouse production.