| Woody plants have important economic, biologicl and social value, however, their multiplication and superior breeding were restricted by the characteristics of poor growth, long growth cycle, and unprecdictable seed production which was mainly depending on environmental change. Plant tissue culture could achieve the rapid, high-efficiency production of nursery stock. On the other hand, as a new technology for the breeding of superior tree species, plant tissue culture has shortened breeding period, and maximized its genetic gain. But there were still a lot of problems which were hard to solve in woody plant tissue culture, especially for superior tree species. In this paper, limber pine (Pinus flexilis James) and Castanopsis hystrix A. DC were taken as objects, the zygotic embryo and tender apical bud of limber pine and the tender stem segment derived from the stump sprouting of C. hystrix A. DC were used as explants, in order to investigate systematically the exsiting problems, their solutions, and mechinisms between them. Finally, the optimal culture methods of different explants during different culture phases were obtained:(1) Surface sterilization:the optimal method for zygotic embryos of limber pine were soaked 20 min in 20%NaClO solution; the tender buds of limber pine were soaked 20 min in 15%NaClO solution; the tender stem segements of C. hystrix A. DC were soaked 13 min in 0.1%HgCl2 solution after storing in ice-water mixture for 1 d.(2) Bud meristem induction:the disinfected zygotic embryos of limber pine were inoculated in GD medium supplemented with BA2.0 mg·L-1, after being cultured in darkness for 2 weeks they were exposed in light for other 2 weeks; the tender buds of limber pine were cultured in SH medium containing BA0.5 mg·L-1 and NAA0.1 mg·L-1 for 4 weeks in light; in order to reduce browning, the tender stem segments of C. hystrix A. DC were inoculated in SH medium containing PVP1.0 g·L-1and BA0.5 mg·L-1, cultured in darkness for 1 week, then exposed to lightness for another 3 weeks. Or inoculated in SH medium supplemented with AC0.5 g·L-1和BA0.5 mg·L-1, the explants were moved to the other side of medium at the 2nd d, and subcultured to the same medium at the 2nd and 4th weeks, respectively, and totally cultured for 6 weeks.(3) Subculture and multiplication of bud meristem:SH medium supplemented with AC0.5 g·L-1, sugar25 g·L-1, agar6 g·L-1, pH was adjusted to 5.2, was suitable for the differentiation of zygotic embryos of limber pine; the adventitious shoots induced from the tender buds of limber pine were elongated in SH basal medium; and the adventitious shoots differentiated form zytotic embryos and the tender buds were multiplicated in SH medium containing BA2.0 mg·L-1, for 4 weeks. Then transferred to SH basal medium or containing AC0.5 g·L-1 for elongation; the tender stem segments of C. hystrix A. DC were subcultured in SH medium supplymented with BA 0.5 mg·L-1, NAA 0.1 mg·L-1 and AC0.1 g·L-1.(4) Rhizogenesis and plantlet transplantation:the adventitious shoots derived from zygotic embryos and the tender buds of limber pine,1.0-1.5 cm in length, were induced for rooting in 1/2GD medium, following 2 weeks cultured in darkness and 4 weeks in light. After being acclimated 1 week in room, the rooted shoots were removed from medium, and transplanted in sterilised substrate (Vpeat: Vvermiculite=1:1), the survival rate was 64.86%。(5) The success of woody plants tissue culture was the consequence of a series of factors co-acting. The ontogenic phase and physiological status of explant was the key of plant tissue culture, medium type and plant growth regulator controled the development direction of explant, and the other additions had an adjustment function on growth.
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