Isolation and regeneration of protoplasts of the moss Physcomitrella patens.

This method is adapted from a protocol described by Grimsley et al. (1977). For more information about P. patens as a model organism, see The Moss Physcomitrella patens: A Novel Model System for Plant Development and Genomic Studies (Cove et al. 2009a). For details about the growth of P. patens on cellophane overlay plates, see Culturing the Moss Physcomitrella patens (Cove et al. 2009b). For protocols describing the use of P. patens protoplasts after they have been isolated, see Somatic Hybridization in the Moss Physcomitrella patens Using PEG-Induced Protoplast Fusion (Cove et al. 2009c), Transformation of the Moss Physcomitrella patens Using Direct DNA Uptake by Protoplasts (Cove et al. 2009d), and Transformation of the Moss Physcomitrella patens Using T-DNA Mutagenesis (Cove et al. 2009e). For details on the use of a hemocytometer to estimate protoplast density, see Estimation of Cell Number by Hemocytometry Counting (Sambrook and Russell 2006).