Rathinapriya Periyasamy, Satish Lakkakula, Rameshkumar Ramakrishnan, Pandian Subramani, Rency Arockiam Sagina, Ramesh Manikandan
1Department of Biotechnology, Science Campus, Alagappa University, Karaikudi, Tamil Nadu 630 003 India.
2Department of Biotechnology Engineering, The Jacob Blaustein Institutes for Desert Research, Ben-Gurion University of Negev, 84105 Beer Sheva, Israel.
Physiol Mol Biol Plants. 2019 Mar;25(2):533-548. doi: 10.1007/s12298-018-0619-z. Epub 2018 Nov 17.
An efficacious, reproducible direct in vitro regeneration system has been developed from leaf base segments (LBs) of six high yielding genotypes of foxtail millet ( (L.) Beauv.). LBs excised from 4-day-old seedling were inoculated on Murashige and Skoog (MS) medium supplemented with different types and concentrations of cytokinins. The shoots induced per explant significantly increased with the supplementation of BAP to auxin containing medium. The results showed that a maximum shoot induction, 58.8% was obtained on MS medium incorporated with 8.9 µM BAP and 2.7 µM NAA in 'CO5' genotype. Further, the highest frequency of multiple shoots was produced on MS(I) medium containing 8.9 µM BAP, 2.7 µM NAA, 700 mg L proline, 0.5 mg L cysteine, 2.0 mg L glycine and 150 mg L arginine. MS(I) medium additionally fortified with 5.0 g L activated charcoal (AC) was found to achieve the best precocious plant regeneration. Elongated shoots were rooted on half-strength MS medium amended with 2.9 µM IAA and achieved maximum root number (8.7) within 10 days. Rooted plantlets were acclimated in soil with 92% survival rate. Molecular marker analysis of in vitro regenerated and field grown plants revealed no somaclonal variations. Briefly, amino acids and activated charcoal could significantly enhance the foxtail millet direct multiple shoot proliferation and plant regeneration. Here we report, a short-term, genotype independent, direct plant regeneration protocol for future genetic transformation studies in foxtail millet genotypes.
已从六种高产基因型谷子（Setaria italica (L.) Beauv.）的叶基部切段（LB）开发出一种高效、可重复的直接体外再生系统。从4日龄幼苗上切下的叶基部切段接种在添加了不同类型和浓度细胞分裂素的Murashige和Skoog（MS）培养基上。在含生长素的培养基中添加BAP后，每个外植体诱导出的芽显著增加。结果表明，在‘CO5’基因型中，添加8.9 μM BAP和2.7 μM NAA的MS培养基上获得了最高的芽诱导率，为58.8%。此外，在含有8.9 μM BAP、2.7 μM NAA、700 mg L脯氨酸、0.5 mg L半胱氨酸、2.0 mg L甘氨酸和150 mg L精氨酸的MS(I)培养基上产生了最高频率的丛生芽。发现额外添加5.0 g L活性炭（AC）的MS(I)培养基能实现最佳的早熟植株再生。伸长的芽在添加2.9 μM IAA的1/2强度MS培养基上生根，并在10天内达到最大根数（8.7）。生根的小植株在土壤中驯化，成活率为92%。对体外再生植株和田间种植植株的分子标记分析表明没有体细胞克隆变异。简而言之，氨基酸和活性炭可以显著提高谷子直接丛生芽增殖和植株再生。在此，我们报告一种短期、不依赖基因型的直接植株再生方案，用于未来谷子基因型的遗传转化研究。
