Improvements of sporophyte culture and comparison of salinity tolerance in three strains of Ceratopteris richardii

Increasing soil salinity on cultivated lands under irrigation is a significant problem confronting world crop production. Due to the multigenic basis of the trait, and problems associated with the regeneration of tolerant plants from tolerant cell cultures, there has been limited progress in the development of salt tolerant crop plants. The use of whole plant mutagenesis and selection for single gene mutants conferring salt tolerance would be useful in studying the genetics associated with the trait. This study documents the selection of salt tolerant mutants in sporophytes of the fern Ceratopteris richardii. In an effort to improve general sporophyte culture, several aspects of in vitro culture were investigated and a comparative study of 'optimized' culture conditions and NaCl tolerance was also performed. In addition, the feasibility of completing the life cycle under in vitro conditions was investigated.

The selection of a mutant (β2-1), conferring tolerance to NaCI was successful. The mutant strain clearly exhibited a higher level of tolerance relative to the parental strain (Nα23-14) and the wild type. Optimized culture conditions significantly enhanced quantitative tolerance on 65 and 80 mM NaCl for each strain tested. This tolerance was transmitted from the Ml to the M2, and from the M2 to the M3 generations, which is strong evidence of a genetic basis. However, because of qualitative variability in tolerance, genetic characterization of the mutant was not possible.

The improvement of general sporophyte culture included investigations of solidifying agents, light intensity, temperature and nutrient media. Both root and shoot growth were significantly enhanced on washed Difco Bacto™ agar. High light intensities (>100 μmol m^-2 s^-1 ) were inhibitory to sporophyte growth. Maximum growth appeared to occur at a temperature of 27.6°C. However, because this study was not replicated, the significant difference seen cannot be unequivocally associated with the variable of temperature. The use of 1/2X DeGreef and Jacob™ nutrient media supported healthy sporophyte growth in both short (18 days) and long term (3 months) cultures when compared to the other medium tested. A comparative study of improved vs traditional culture conditions indicated the improved conditions significantly enhanced sporophyte growth. Treatment with 5.0 X 10^-3 and 10^-2M filter sterilized GA₃ accelerated sporangial development. However, efforts to complete the life cycle under in vitro conditions were unsuccessful.