The physiology of somatic embryogenesis in carrot (Daucus carota L.) and cotton (Gossypium klotzschianum Anderss.)

Abstract

Systems were developed to obtain high frequency of somatic embryogenesis in carrot (Daucus carota L.) and cotton (Gossypium klotzschianum Anderss.). Wild carrot was far superior to cultivated carrot for the establishment of embryogenic cultures. Calli which consistently gave rise to large numbers of somatic embryos were characteristically smooth and yellow. In order to maintain suspension cultures which consisted entirely of proembryonic masses (PEMs), the cultures were initially filtered through a 52 (mu)m screen. Proembryonic masses, which passed through the screen, were separated from nonembryogenic clumps and cells by density centrifugation in a Ficoll gradient. The PEMs developed into embryos once the 2,4-D was removed from the medium. Embryogenic cotton callus was more yellow, slower-growing, and more nodulated than nonembryogenic callus. The quantity of somatic embryos produced from embryogenic cotton callus was dependent on light intensity, callus source, growth phase of the callus used for the initiation of the suspension cultures, concentration of 2,4-D in liquid culture, and incubation period in that 2,4-D-containing medium. The quality of somatic embryos was influenced by the length of incubation in the 2,4-D-containing medium and the frequency of washes with auxin-free medium after 2,4-D removal. Plant regeneration from cotton somatic embryos was dependent on the physiological and genetic state of those embryos, the carbohydrate source, and the physical state of that medium. Change in specific activity of nitrate reductase and malate dehydrogenase appear to be associated with growth phases of cultures in general and are not specifically linked to embryogenesis. Cotton embryos which were placed on media containing either lactose, glucose, or sucrose contained similar levels of acid phosphatase, malate dehydrogenase, and peroxidase after 2 weeks in culture, and equal amounts of indoleacetic acid after 8 weeks in culture. Morphologically, these embryos developed differently; embryos of glucose and sucrose formed callus, and embryos on lactose underwent further organized development.