Roberts, Alison, W
Cellulose Synthase-like D (CSLD) genes; Physcomitrella patens; tip growth; fluorescence microscopy; green fluorescent protein (GFP); targeted gene deletion
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Physcomitrella patens is a non-vascular plant with a relatively small genome and is amongst the few eukaryotic organisms that have a high rate of homologous recombination. This is valuable in biological research because it allows for targeted genetic modification of the organism. In vascular plants like Arabidopsis thaliana, a model organism, Cellulose Synthase-like D (CSLD) genes have been discovered to be important in tip growth. This type of growth is observed in the pollen tubes and root hairs of these plant types. The CSLD genes in Arabidopsis were found to play a crucial role in the growth of root hairs and the production of cellulose or cellulose-like β-1,4-glucan chains in root hair tips. The CSLD genes have also been recognized to be important in pollen tube growth of vascular plants. Physcomitrella patens also contains genes similar to the vascular plant CSLDs, but their functions are not yet fully understood. Within the P. patens genome there are eight genes that make up the CSLD gene family. Additionally, the life cycle of P. patens includes a stage that consists primarily of tip growing cells. This growth stage can be optimized in order to study the role of CSLD genes in tip growth of P. patens.
In an effort to further study the roles of the CSLD genes in tip growth of P. patens, we constructed a plasmid that expresses the CSLD1 protein with a green fluorescent protein (GFP) tag. This allowed us to visualize the expression of CSLD1 in living cells using fluorescence microscopy. We also constructed plasmids that were designed to remove specific CSLD genes from the genome and transformed them into wild type or CLSD1 knockout tissue of P. patens. This created single or double knockout mutants that could then be compared to the wild type for changes in the phenotypic characteristics of the plant. These findings will aid in uncovering the roles of the CSLD gene family in P. patens and may provide insight into the functions of these genes in other plants.