烟草种子直接诱导愈伤组织的方法探究

Research on the method of directly inducing callus from tobacco seeds

以烟草种子为材料,探究从种子直接诱导出愈伤组织的方法。结果表明:烟草种子愈伤诱导受2,4-二氯苯氧乙酸(2,4-D)和6-苄氨基嘌呤(6-BA)不同激素配比影响,其中,在0.1~2.0 mg/L 2,4-D和6-BA的激素浓度范围内均能诱导出愈伤组织,但大小和质量各不相同。值得注意的是,种子在含1.0 mg/L 2,4-D+1.0 mg/L 6-BA的培养基中培养,能较快诱导出愈伤组织,但愈伤组织块较为松散;而在含0.1 mg/L 2,4-D+1.0 mg/L 6-BA的培养基中培养,诱导出的愈伤组织较为紧密。进一步通过石蜡切片观察发现,在1.0 mg/L 2,4-D+1.0 mg/L 6-BA培养条件下诱导出的愈伤组织细胞较大,但排列不规则,多为非胚性细胞;相反,在0.1 mg/L 2,4-D+1.0 mg/L 6-BA培养条件下诱导出的愈伤组织细胞虽然较小,但排列规则,多为胚性细胞。研究结果为从烟草种子诱导培养愈伤组织提供新的技术参考,也为烟草遗传转化研究提供便利。

Tobacco seeds were used as explants to explore how to directly induce callus.The results showed that the callus induction of tobacco seeds was affected by different hormone ratios of 2,4-dichlorophe noxyacetic acid(2,4-D)and 6-benzylami nopurine(6-BA),and the combined use of hormones 2,4-D and 6-BA in the concentration range of 0.1-2.0 mg/L could induce callus,but different in size and quality.It was worth noting that when the seeds were cultured on the medium containing 1.0 mg/L 2,4-D+1.0 mg/L 6-BA,callus was rapidly induced,but the callus mass was relatively loose.In comparison,the callus was more compact when the seeds were cultured on the medium containing 0.1 mg/L 2,4-D+1.0 mg/L 6-BA.Moreover,paraffin sections showed that the callus cells induced by the hormone combination of 1.0 mg/L 2,4-D+1.0 mg/L 6-BA were larger but irregularly arranged and most of them were non-embryonic cells.On the contrary,the callus cells induced by the hormone combination of 0.1 mg/L 2,4-D+1.0 mg/L 6-BA were small and regularly arranged,and most of them were embryonic cells.The results of this study provide new technical support for the induction of callus from tobacco seeds and facilitate the research on genetic transformation in tobacco.