Slow desiccation leads to high-frequency shoot recovery from transformed somatic embryos of cotton (Gossypium hirsutum L. cv. Coker 310 FR).

In Agrobacterium-mediated genetic transformation of cotton (Gossypium hirsutum L. cv. Coker 310FR) the frequency at which somatic embryos were converted to plantlets was significantly improved by subjecting the embryos to slow physical desiccation. We used Agrobacterium strain GV3101 containing the binary vector pGSFR with the nos-nptII gene for in vitro selection and the 35S gus-int fragment as a reporter to optimize the transformation protocol. Although the concentration of kanamycin was reduced during embryogenesis and embryo maturation, even at the lower levels somatic embryos were predominantly abnormal, showing hypertrophy and reduced or fused cotyledons or poor radicle ends. A majority of these embryos (more than 75%) were beta-glucuronidase (GUS)-positive. Embryos with an abnormal appearance showed a very poor conversion to plantlets. However, these embryos, when subjected to slow physical desiccation followed by transfer to fresh medium, regenerated single or multiple shoots from the cotyledonary end. These shoots could be grafted on wild-type seedling stocks in vitro, which, following their transfer to soil, developed normally and set seeds. Regenerated plants tested positive for the transgene by Southern analysis. An overall scheme for the high-frequency production of cotton transgenics from both normal and abnormal appearing somatic embryos is presented.

Totipotency of coleoptile tissue in indica rice (Oryza sativa L. cv. ch 1039).

Embryogenic and non-embryogenic calluses were induced from 3,4,5 and 7d old coleoptile segments of indica rice (Oryza sativa L. cv. CH 1039). Compact, globular, yellow and creamy embryogenic and white friable non-embryogenic callus arose from the cut end and entire length of the coleoptile segments. Murashige and Skoog's (MS) medium supplemented with 2.5mg/1 2,4-D was used as callus induction medium. Plant regeneration from coleoptile segments occurred with the transfer of embryogenic callus to MS basal medium supplemented with 2.0mg/1 BAP and 0.5mg/1 NAA in combination. Average number of regenerated plants from one coleoptile ranged from9.1 to 14.0.Four day old coleoptiles showed the highest frequency of plant regeneration.