Particle bombardment mediated transformation and GFP expression in the moss Physcomitrella patens

There are few plants facilitated for the study of development, morphogenesi s and gene expression at the cellular level. The moss Physcomitrella patens can be a very useful plant with several advantages: simple life cycle cont aining a major haploid gametophyte stage, easy manipulation, small genome s ize (6 x 10(8) bp) and high similarities with higher plants. To establish t he transformation system of mosses as a model for basic plant research, a s eries of experiments were performed, Mosses were cultured in cellophane ove rlaid BCD media, transformed by particle bombardment and selected by the ch oice of appropriate antibiotics. Initial transformants appeared 8 d or 14 d after selection, showing different sensitivities toward the antibiotics us ed, Heat treatment during the preparation of particles revealed that denatu ring the DNA enabled a more efficient way to deliver a transgene into the c hromosome. This was proven by the increase in the number of transformants b y five times in the plants with denatured DNA, In the test for the repairin g capacity of mosses, 154 and 195 transformants survived from Id and 3d inc ubations, respectively, indicating that a longer period of incubation seeme d to be recommendable for better survival. The selected transformants were further analyzed at the DNA and expression level. Transformed genes were co nfirmed by PCR where all the transformants showed the expected size of ampl ification. Histochemical beta-glucuronidase (GUS) and green fluorescent pro tein (GFP) expression also confirmed the integration of exogenous DNA, In a comparison of the two different forms of GFP, soluble-modified GFP (smGFP) expressed stronger signals than modified GFP (mGFP) due to its improved so lubility. Confirmation of the transgene in the chloroplast transformation h as improved the applicability of moss as a model system for the study of ba sic biological researches.