大麦茎尖愈伤组织的诱导和植株高频再生

为明确大麦茎尖高频再生培养体系的影响因素,以大麦品种甘啤3号、甘啤4号和甘啤6号为材料,研究颖壳去除方法、灭菌处理方式、无菌苗的苗龄、茎尖切段的大小和位置以及外源激素(2,4-D)浓度对大麦茎尖愈伤组织培养和植株高频再生的影响。结果表明,大麦种子颖壳去除的最佳方法是将种子浸泡在30%的硫酸中,并置于水平摇床30℃、180r·min~(-1)摇动1h;同时发现参试品种中甘啤4号发芽率最高;75%的乙醇1.5min+2%的NaClO 15min的灭菌处理效果最好,其带菌率和发芽率与其他处理有显著差异,并且对种子伤害较小,幼苗整齐度较好,有利于无菌苗的培养;茎尖外植体愈伤组织的形成和植株再生频率与茎尖切段的位置和苗龄密切相关,3d苗龄、茎尖基部2mm处的切段效果最好;3mg·L~(-1) 2,4-D适宜愈伤组织的离体培养,在出愈率和植株再生率上差异显著,可以诱导出大量愈伤组织。

Callus Induction and High Frequency Regeneration of Plant from Shoot Tip of Barley (Hordeum vulgare L.)

High frequency regeneration system for tissue culture possesses high efficient and stable regeneration,and it is also the foundation of genetic transformation.Explant materials of different parts were tried to establish high frequency regeneration system in barley.In order to study the factors related to high frequency regeneration from shoot tip of barley,Ganpi 3,Ganpi 4 and Ganpi 6 were used to study the effects of the dehulling methods,sterilization ways,seedling age,shoot tip size and location,and the concentration of exogenous hormone (2,4-D) on the stem tip callus induction and the plant high frequency regeneration.The results showed that the optimal dehulling way is to use 30％sulfate and shake for an hour at 180 r·min-1 and 30℃,and Ganpi 4 has the highest germination rate.The optimal method for surface sterilization was soaking in 75％ ethanol for 1.5 min and then 2％ sodium hypochlorite for 15 min,which is benefit for the aseptic seedling cultivation,with significant difference in microorganism-carrying and germination rate for seeds,less damage on seed and the good uniformity for seedlings.Callus formation of the shoot tip explants and plant regeneration frequency are closely related to shoot tip position and seedling age,and the optimal length of basal segments is 2 mm and the optimal seedling age is 3 d.The optimal concentration of 2,4-D for callus culture is 3 mg·L-1,which indicated the most significant difference in recovery and differentiation rates and could induce a large number of callus.