Ebrahimie Esmaeil, Hosseinzadeh Abdolhadi, Nagavi Mohammad Reza, Ghannadha Mohammad Reza, Mohammadie-Dehcheshmeh Manijeh
Department of Agronomy and Plant Breeding, Faculty of Agriculture, University of Tehran, Karaj 31587-11167, Iran.
Pak J Biol Sci. 2007 May 1;10(9):1360-70. doi: 10.3923/pjbs.2007.1360.1370.
Rapid and genotype-independent protocols for two direct in vitro morphogenesis pathways including direct shoot organogenesis from embryo and direct shoot proliferation from node have been developed in cumin (Cuminum cyminum L.). Direct regenerations occurring without passing callus phase are important since fewer somaclonal variation and genotype-dependency are likely to arise from these methods in comparison with regenerations trough callus. After embryo culture, shoots with single-cellular origin were regenerated from the meristematic zone of embryo without any intermediate callus phase. In contrast, proliferated shoots with multi-cellular origin were directly regenerated from the axillary buds (meristems) of node explants. Effects of different concentrations of 6-Benzylaminopurine (BAP), alpha-Naphthaleneacetic Acid (NAA) and Indole-3-kcetic Acid (IAA) on B5 medium of embryo and node cultures as well as subculture were studied in detail. In direct organogenesis pathway from embryo explant, 0.1 mg L(-1) NAA + 1 mg L(-1) IAA resulted the highest shoot regeneration response (89.5 shoots per regenerated explant), whereas 0.1 mg L(-1) BAP + 1 mg L(-1) NAA was the most effective combination in direct shoot proliferation from node explant (42 shoots per regenerated explant). BAP (cytokinin) revealed the inhibitory effect on induction of direct shoot organogenesis pathway from embryo explant, while low concentration of BAP (0.1 mg L(-1)) had positive effect on direct shoot proliferation pathway from node explant. Subculturing was not necessary for shoot multiplication and elongation in embryo culture, whereas multiplication and elongation of shoots in node culture were associated to subculture on growth regulator-free medium. In other part of study, the behavior of different cumin genotypes in direct regeneration pathways was studied. Both direct organogenesis and direct proliferation pathways were applicable to different cumin genotypes and regenerated plants were phenotypically normal. This study supports the feasibility of combined direct regenerations protocols from embryo and node of cumin in germplasm conservation by in vitro cloning and genetic improvement programs.
孜然（Cuminum cyminum L.）已开发出快速且不依赖基因型的两种直接体外形态发生途径的方案，包括从胚直接诱导芽器官发生和从节直接诱导芽增殖。直接再生不经过愈伤组织阶段很重要，因为与通过愈伤组织再生相比，这些方法产生体细胞无性系变异和基因型依赖性的可能性较小。胚培养后，从胚的分生组织区再生出单细胞起源的芽，无任何中间愈伤组织阶段。相反，多细胞起源的增殖芽直接从节外植体的腋芽（分生组织）再生。详细研究了不同浓度的6-苄基腺嘌呤（BAP）、α-萘乙酸（NAA）和吲哚-3-乙酸（IAA）对胚和节培养以及继代培养的B5培养基的影响。在从胚外植体的直接器官发生途径中，0.1 mg L(-1) NAA + 1 mg L(-1) IAA产生了最高的芽再生响应（每个再生外植体89.5个芽），而0.1 mg L(-1) BAP + 1 mg L(-1) NAA是从节外植体直接诱导芽增殖中最有效的组合（每个再生外植体42个芽）。BAP（细胞分裂素）对从胚外植体诱导直接芽器官发生途径有抑制作用，而低浓度的BAP（0.1 mg L(-1)）对从节外植体直接诱导芽增殖途径有积极作用。胚培养中芽的增殖和伸长不需要继代培养，而节培养中芽的增殖和伸长与在无生长调节剂的培养基上继代培养有关。在研究的其他部分，研究了不同孜然基因型在直接再生途径中的表现。直接器官发生和直接增殖途径都适用于不同的孜然基因型，再生植株表型正常。本研究支持了通过体外克隆和遗传改良计划，将孜然胚和节的直接再生方案相结合用于种质保存的可行性。
