Mise au point d'une technique de culture in vitro d'embryons immatures de Phaseolus

Development of an in vitro culture technique for immature Phaseolus embryos. In the interspecific crosses Phaseolus polyanthus (or P. coccineus) (i?) x P. vulgaris, the hybrid embryos abort very early. Therefore, it is essentiel to develop an in vitro culture technique that allows the rescue of beau embryos at globular or early heart-shaped stages. After several trials conceming the salts composition, the sugar rate and the amino acid concentration of différent in vitro culture media, a technique has been developed for heart-shaped Phaseolus embryos. This technique consists of two stages. In a first step, embryos are cultivated under darkness until their germination on a medium containing the salts of Gamborg et al. (1968), 400 mg . 1` (5mM - 1 -') NHNO,, 1 mg . 1-' thiamine HCI, 5 mg . l` nicotinic acid, 0.5 mg - l` pyridoxine, 1,000 mg . l` -glutamine, 1,000 mg . l` casein hydrolysate, 100 mg . l` myo-inositol, 0.028 mg . P N6-benzyladenine, 30 g . l` sucrase, and 8 g -1-' DIFCO agar. After germination, the embryos are cultivated under light on a second medium that does not contain any NHNO, complément and is poorer in amino acids (100 mg 1-' L-glutamine). Developed with six deys old heart-shaped embryos of the P. vulgaris Bico de Ouro (NI 637) variety, this technique has proved its efficiency with other P. vulgaris and P. polyanthus génotypes. It allows an average régénération rate of 30% from the total number of cultivated embryos