蛇足石杉侧芽的发生与培养研究

Study on lateral buds induction and culture of Huperzia serrata

目的蛇足石杉Huperzia serrata自然繁殖能力低,资源极其匮乏,对蛇足石杉的快速繁殖技术进行探讨。方法取活体茎尖,依次用多种消毒剂进行表面消毒,再用孔雀石绿及原子吸收光谱（AAS）联合杀灭内生真菌,得到无菌苗接种到1/2MS＋25 g/L蔗糖的培养基中,无菌苗在二叉分枝后的新枝长到4-5 cm时剪下进行继代培养。结果无菌苗分别在4次二叉分枝生长继代培养后,得到了数量不等的侧芽,侧芽的发生位置位于茎段中部或上部近茎尖处,侧芽的结构与芽孢的结构明显不同;不同植株培养获得的侧芽数不同,最多的植株可产生14个侧芽,一个侧芽发生点可产生一个或多个侧芽;去除茎尖顶端优势后,茎段中部的侧芽生长速度加快;不同高度的侧芽拨离母株后,成活率差别大,5 mm以上的侧芽接种到1/2 MS＋30 g/L蔗糖的培养基上,存活率可达90%,120 d后生长为2-3 cm的幼苗,低于5 mm的侧芽,拨离接种后死亡率较高。结论蛇足石杉在离体培养中,可打破自然生长状态下叶腋内只产生孢子囊而不产生侧芽的生长方式,可形成多个增殖侧芽,侧芽剥离后极易产生不定根,为蛇足石杉的试管苗增殖方式提供重要参考。

Objective Due to the low reproductive capacity and scarce resources of Huperzia serrata, and in vitro culture of H. serrata has not been really successful, we wish to make a good progress in rapid propagation in this study. Methods Living shoot tips were used as the materials, after a variety of surface sterilization, malachite green and AAS were used to eliminate the endophytic fungi of the seedling. Then, the aseptic seedlings were transplanted to the medium with 1/2 MS ＋ 25 g/L sucrose to be cultured. After the new dichotomous branching occurred, and the branches grew to 4—5 cm high, we cut it and subcultured for the new dichotomous branching. Results After four times of subculture of dichotomous branching, the aseptic seedlings got the different number of proliferating lateral buds, which came from the middle or top of the stem. Comparing the structure of lateral buds with the gemma, we found it was significantly different from the gemma. The number of lateral buds from different individuals was different, the most lateral buds could get to 14 in one seedling, some lateral bud generating point can produce two or more lateral buds. After removing the apical dominance of stem tip, the lateral buds in the middle of the stem grew faster. The surviving rate of different height lateral buds was different when they separated from the stock plant. Lateral buds which beyond 5 mm, the surviving rate could get 90%, while cultured in medium of 1/2 MS ＋ 30 g/L sucrose, after 120 d, it grew into seedling about 2—3 cm high, lateral buds less 5 mm had high mortality with the same cultured medium. Conclusion Though in vitro culture, H. serrata can break the reproductive way by which the leaf axil could generate spore only but no lateral bud naturally, and can produce the multiple lateral bud proliferation. The lateral buds can easily produce adventitious roots by stripping, which could provide an important view for rapid reproduction of H. serrata.