RECOVERY OF HAPLOID PLANTS FROM ASPARAGUS MICROSPORE CULTURE

Asparagus (Asparagus officinalis L.) microspore culture was performed in an array of experiments that assessed the roles of plant growth and culture conditions. The following protocol provided the best results. Flowers with microspores at the late uninucleate stage of development were collected from greenhouse plants grown at 22:18 degrees C (light :dark) and stored at 5 degrees C for 3 days. One millilitre of MS medi um plus 0.2 g/L yeast extract, 500 mg/L casein hydrolysate, 800 mg/L g lutamine, 2.0 mg/L naphthaleneacetic acid, 1.0 mg/L benzyladenine, and 6% sucrose (MSFY) was conditioned with 10 anthers/ml for 1 week, afte r which it was filtered. One hundred anthers were added to shed their microspores (1.6 x 10(5) per mt) and were removed after 3 weeks when 0 .5 mL of fresh medium was added. Cultures were incubated at 35 degrees C for 1 week, then 30 degrees C for 5 weeks. Microcalli were collecte d subsequently on a 100-mu m screen and placed on induction medium (MS FY minus yeast extract, plus 3 g/L gelrite) in darkness at 35 degrees C for 4 weeks and then in light at 25 degrees C for 4 weeks. Shoots, r oots, and bipolar embryos were produced. The latter were transferred t o maturation medium (MS plus 0.1 mg/L naphthaleneacetic acid, 0.5 mg/L kinetin, 3% sucrose, 3 g/L gelrite, and 0.65 mg/L ancymidol) for 4 we eks, then to germination medium (MS plus 1.0 mg/L gibberellic acid, 3% sucrose, 3 mg/L gelrite). Plantlets were grown and maintained on matu ration medium. Approximately 0.3% of the cultured microspores produced calli, and 85% of calli produced plantlets. Of 10 plants analyzed, 2 were haploid, 7 were diploid and, 1 was tetraploid.