Transgene Expression In Specific Neurons Of C.elegans

By contrast with other model animals, nervous system of C. elegans is simple. There are only 302 neurons in hermaphrodite and have been classified into several groups--namely, motorneuron, sensory neuron, interneuron and so on. Though less those neurons, they generate such complicated connections which carry out significantly indispensible functions according to the developmental orders. The subtle connections between neurons approximately contain 5000 chemical synapses, 2000 neuromuscular junctions and 600 gap junctions. At some extent, neurons and synaptic linkages have already been studied by serials of electron microscopic section techniques in the development.Because of its simple structure and sequenced genome, C. elegans have already used as model organism for scientific researches. In order to show how valuable C. elegans is and to offer more available advantages to researchers, the main idea of my topic: firstly, is going to mark the specific location of C. elegans neurons; secondly, we can preferably study the detailed functions of those neurons based on which we have located. Although a myriad of techniques have been sufficiently utilized in the area of transgene expression in some specific tissues, very specific promoter is limited so that we can't dependent on the only method. In the term of disadvantage, we choose two site-specific recombination system (FLP/FRT, Gateway) and confocal laser scanning technique to locate single neuron, and obtain numerous images. Since most of promoters have intersection when express different fluorescent protein, we sign neurons by different colors for the sake of distinguishing some expressions promoted by different promoters. After locating target neuron, other translated proteins such as ChR2 or NPHR, GCaMP and CED-3 might be used in further study concerning neuron functions of nervous system of C. elegans.Many in vitro and in vivo applications for transgenesis require co-expression of heterologous genes. The use of second spliced leader (SL2) in multicistronic expression vectors enables the expression of many genes controlled by one promoter in target cells or the whole organisms. Here we choose SL2 to express multiple cistronics under specific promoter, based on mocular tecnique and confocal laser scanning technique. As a result, we have a conclusion that mutiple tandem-SL2 does not affect other gene expression; the fluorescence output of GFP is dependent on the number of SL2-GFP repeats in each construct and on their translation efficiency but not on different genomic integration efficiency. SL2-dependent polycistronic gene expression opens up the possibility, to titrate the protein expression level by expressing the gene of interest under the optimal SL2-tandem repeats.