Abstract
Here we describe a rapid high-throughput method for performing RNA interference (RNAi) in moss, in which phenotyping is performed within 1 week after transformation. The moss Physcomitrella patens is a great plant model system for reverse genetic studies due to its amenability to homologous recombination as well as RNAi. Our lab has developed a rapid RNAi assay to screen for growth phenotypes in moss protonemal tissue. Here we describe how we have recently further facilitated this assay by modifying the PEG-mediated transformation protocol allowing for transformations to be carried out in a semiautomated fashion in a 96-well plate format.
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Acknowledgments
We thank Lawrence Winship for building the temperature control systems. We thank Paula Franco and Luis Vidali for initiating the project and David O’Donnell for performing pilot screens. We are grateful to members of the Bezanilla, Baskin and Hepler labs for helpful suggestions and recommendations. This work was supported by a fellowship from the David and Lucille Packard Foundation to (MB) and the University of Massachusetts – Amherst Plant Biology Graduate Program (SW).
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Wu, SZ., Bezanilla, M. (2012). Transient RNAi Assay in 96-Well Plate Format Facilitates High-Throughput Gene Function Studies in Planta. In: Normanly, J. (eds) High-Throughput Phenotyping in Plants. Methods in Molecular Biology, vol 918. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-995-2_17
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DOI: https://doi.org/10.1007/978-1-61779-995-2_17
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Publisher Name: Humana Press, Totowa, NJ
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