An efficient in vitro protocol for high-frequency polyploidization for the first time in gerbera hybrid (BGC-2019-01) was developed in the present study. Two-week-old in vitro-developed shoots (tips) were treated individually with 0.1%, 0.25% and 0.5% (w/v) colchicine solutions for 4, 6, 8, and 12 h. The colchicine-treated shoot tips were then inoculated on Murashige and Skoog (MS) medium fortified with 1.5 mg/l meta-Topolin for multiple shoot proliferation and later transferred into 1.5 mg/l indole-3-acetic acid-fortified MS medium for rooting of shoots. The ploidy levels of the colchicine-treated and regenerated plantlets along with the non-treated ones were confirmed via flow cytometry analysis and metaphasic chromosome count. The highest frequency of tetraploid plantlets (50%) were obtained when shoot tips were treated with 0.1% colchicine for 4 h. Morphological observations revealed that induced tetraploid plantlets exhibited delayed fresh shoot initiation, fewer but longer shoots, as well as fewer but broader leaves. Likewise, the study of stomata revealed that in comparison to their diploid counterparts, the tetraploid plantlets exhibited less frequent yet significantly larger stomata, and higher number of chloroplasts. The tetraploids were recorded with significantly higher chlorophyll, carotenoid, and anthocyanin content during the photosynthetic pigment analyses. During ex vitro acclimatization and field growth, the tetraploid plants exhibited delayed proliferation but with higher vigor and thickened broad leaves. The genetic uniformity among the diploid and the tetraploid plants was confirmed using conserved DNA-derived polymorphism (CDDP), directed amplification of minisatellite-region DNA (DAMD), inter simple sequence repeats (ISSR), and start codon targeted (SCoT) polymorphism marker systems. The tetraploids developed in the present study would be of immense importance for the genetic improvement of gerbera as far as its ornamental values are concerned.

本研究首次在非洲菊杂交种 (BGC-2019-01) 中开发了一种有效的体外高频多倍化方案。两周大的体外发育芽（尖端）分别用 0.1%、0.25% 和 0.5%（ w / v ）秋水仙碱溶液处理 4、6、8 和 12 小时。然后将经秋水仙碱处理的茎尖接种到用 1.5 mg/l meta -Topolin 强化的 Murashige 和 Skoog (MS) 培养基上，以进行多枝增殖，然后转移到 1.5 mg/l 吲哚-3-乙酸强化的 MS 培养基中用于生根的芽。通过流式细胞术分析和中期染色体计数确认了经秋水仙碱处理和再生的植株以及未经处理的植株的倍性水平。当茎尖用0.1%秋水仙碱处理4小时时，获得四倍体植株的最高频率（50%）。形态学观察表明，诱导的四倍体幼苗表现出新芽萌发延迟，芽较少但较长，以及叶片较少但较宽。同样，气孔研究表明，与二倍体对照相比，四倍体幼苗表现出频率较低但明显较大的气孔和叶绿体数量。在光合色素分析过程中，四倍体的叶绿素、类胡萝卜素和花青素含量显着升高。在离体驯化和田间生长过程中，四倍体植物表现出增殖延迟，但具有较高的活力和增厚的阔叶。 使用保守 DNA 衍生多态性 (CDDP)、小卫星区 DNA 定向扩增 (DAMD)、简单序列重复之间 (ISSR) 和起始密码子靶向 (SCoT) 多态性标记确认二倍体和四倍体植物之间的遗传一致性系统。就其观赏价值而言，本研究中开发的四倍体对于非洲菊的遗传改良具有极其重要的意义。