| During the cultures of immature embryos of wheat (Triticum aestivum L.), some factors, including exogenous phytohormones, surgar concentrations in medium, age of immature embryos and genotypes of donor wheat cultivars(or lines), which may influence the frequency of somatic clones indnction, have been researched. And by counting the regeneration frequency, investigating the degrees of embryogenic maintenance, determing the activities of sugarase and peroxidase (POD) and observing the photographs of Scanning Electronic Microscope (SEM) and of cellular histochemistiy, the changes of embryogenic callus during subcultures and the differences between embryogenic and nonembryogenic calli, have been acquired. The results are the following.1 .Exogenous phytohormones have the significant effects on the somatic clones of immature embryos in wheat. Concentration of 2,4-D in media significantly affects the induction of embryogenic calli derived from immature embryos of wheat, with different frequency between the three genotypes. The most appropriate concentration of 2,4-D is around 2.O----3.Omgl, at which the increase of callus in fresh weight and the percentage of embryogenic callus among the three genotypes are the highest. This capability of 2,4-D can be enforced by the addition of KT or 6-BA in medium. But the enforcement differs between KT and 6-BA. KT has a remarkable role in the induction of embryogenic calli at a relatively higher concentration while 6-BA works well at a lower concentration.2.Concentrations of sugar in media notablely affected the establishment of the somatic clones of immature embryos at about 6.O%(w/v) or 6OglL and at this concentration the biomass of callus and the percentage of embryogenic calli are the highest. Furthxnore, if the concentration is too high or too low, the growth of callus or the percentage of embryogenic calli induction will be affected, even be inhibited.3.Different ages of immature embryos, i. e. developmental days of embryo after pollination, affected the frequency of calli induced and the percentage of embryogenic calli induced among three genotypes of wheat immature embryos. The best period for callus induction differs among different genotypes, and in Yangling may be around 14d after pollination for winter wheat.4.Different genotypes had little effect on the frequency of callus induction but significantly affected the percentage of embryogenic callus induction. Among the 12 cultivars or lines of wheat, every one had higher frequency of callus induction (>90%), while the percentage of embryogenic callus of each differs significantly. Three cultivars, Qianjinzao, Xiao yan NO. 22 and Xinong NO. 85, have higher percentage of embryogeniccallus than the others.5.During subculture, embryogenic callus of most cultivars or lines wnderwent descending trend in regenerability of plantlet, extension of the embryogenical maintenance and the activities of surgarase (transformase) and peroxidase (POD). But there are still cultivars, including Qianjinzao and Xiaoyan NO. 22, which had the stable subcé»®turing characteristics. Furthermore, for these stable cultivars. some different stages of somatic embryos or embryoid can be seen under Scanning Electronic Microscope (SEM) or optic microscope by double staining of histochemical method.6.Between embryogenic callus (EC) and nonembryogenic callus (NEC), there exsited wide-ranging differences, including appcarances, characteristics of physiology and biocheniisty, morphological structure under SEM or microscope. Generally speaking, EC has its granular and compaet appearance with yellowrish or light yellow colour, grows more slowly than NEC, and its regenerability is higher than NEC.Moreover, the metabolism of EC is active. Sometimes there are different stages of embryoid observed under SEM or optic microscope. On paraffin slice stained with safranin and fast green, embryogenic cells appeared round shape, smaller dimension, orderly arrayal, and had larger nuclei, thick cytoplasm, and cellulosized cell... |
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