is a powerful model system to study plant gene function. Unique among plants, moss performs efficient homologous recombination enabling precise genetic manipulation, including gene deletions and allele replacements. With a completely sequenced genome, facile genetic alteration, and a relatively simple developmental pattern, moss provides an unparalleled tool for dissecting plant gene function. Moss is simple to propagate and transform, and requires little space to maintain. This lab manual contains information about propagation, transformation, and DNA and protein isolation. By removing the cell walls from moss tissue, it is possible to generate a suspension of single cells called protoplasts. Moss protoplasts can be transformed with DNA and then regenerated into whole plants. For details on this procedure, please refer to the moss manual. Moss protoplasts given the proper growth conditions will within a few days regenerate into a whole plant. Thousands of plants can be regenerated. Taking pictures of the plants, it is possible to get measurements of overall plant size and shape. Moss performs RNAi-induced gene silencing. Double stranded RNA can be generated by transformation of DNA constructs containing inverted repeats of the sequence targeted for gene silencing. My laboratory uses a robust RNAi assay that enables rapid identification of gene function. Complementation studies provide validation of loss-of-function studies and mechanistic insights into gene function. By targeting untranslated regions of sequence, RNAi-induced phenotypes can be rescued by expression of a construct containing only the open reading frame. For expression of your favorite protein using the maize ubiquitin promoter. Can be used for transient overexpression or for generating a stable line. All vectors have moss sequences for homologous recombination based targeting. When exogenous sequences are inserted into these loci, there is no affect on growth or development. Cloning into these vectors is mediated by Gateway recombination based cloning from Invitrogen. http://www.invitrogen.com/site/us/en/home/Products-and-Services/Applications/Cloning/Gateway-Cloning/GatewayC-Misc/MultiSite-Gateway-Pro-Kits.html Source.
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