金冠苹果成熟胚离体诱导四倍体的研究

 以自然授粉的‘金冠’苹果种子为材料，研究了秋水仙素处理成熟胚离体诱导四倍体技术和嵌合体分离方法。结果表明：将完整胚在0.5%秋水仙素+1%二甲基亚砜的混合溶液中暗处理48 h，然后接种在MS+TDZ 1.0 mg·L^-1+IBA 0.5 mg·L^-1+蔗糖30 g·L^-1+琼脂6 g·L^-1的再生培养基中，变异率高达10.6%。与二倍体相比，四倍体试管苗生长缓慢，节间变短，叶片增大，叶形指数趋小化，气孔密度降低，保卫细胞增大。经压片法鉴定茎尖染色体数和流式细胞仪分析DNA含量，表明多数变异植株为同质四倍体。利用叶片离体再生器官技术，从诱导获得的嵌合体植株中分离出同质突变体。

In Vitro Induction of Tetraploid from Mature Embryos of 'Golden Delicious' Apple

In vitro induction of tetraploid in apple and the technique of chimera separation were studied by treating open pollinated mature seeds of Golden Delicious with colchicine. The most efficient condition for inducing tetraploids seemed to be treating embryos with the solution including of 0.5% colchicine and 1% dimethyl sulfone in darkness for 48 hours, and then inoculation on MS medium supplemented with TDZ 1.0 mg·L^-1, IBA 0.5 mg·L^-1, sucrose 30 g·L^-1 and agar 6 g·L^-1, with 10.6% mutation rate. Compared with the diploid, the tetraploid plants grew slowly, with shorter internode, larger leaves with larger guard cell and lower density of stomata, and smaller leaf shape index. Verification of chromosome number in shoot tip cells by pressing cover slip method and detection of relative nuclear DNA content by flow cytometry proved that most mutated plantlets are homogeneous tetraploid. It was an effective method of separating chimeras into their homogeneous ploidy by in vitro regeneration plantlets from leaf pieces.