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Abstract
背景回顾：Plant genome editing and propagation are important tools in crop breeding and production. Both rely heavily on the development of efficient in vitro plant regeneration systems. Two prominent regeneration systems that are widely employed in crop production are somatic embryogenesis (SE) and de novo shoot regeneration. In many of the protocols for SE or shoot regeneration, explants are treated with the synthetic auxin analog 2,4-dichlorophenoxyacetic acid (2,4-D), since natural auxins, such as indole-3-acetic acid (IAA) or 4-chloroindole-3-acetic acid (4-Cl-IAA), are less effective or even fail to induce regeneration.
研究假设：Based on previous reports that 2,4-D, compared to endogenous auxins, is not effectively exported from plant cells, we investigated whether efflux inhibition of endogenous auxins could convert these auxins into efficient inducers of SE in Arabidopsis immature zygotic embryos (IZEs).
结果1-生长素外排抑制：We show that natural auxins and synthetic analogs thereof become efficient inducers of SE when their efflux is transiently inhibited by co-application of the auxin transport inhibitor naphthylphthalamic acid (NPA). Moreover, IZEs of auxin efflux mutants pin2 or abcb1 abcb19 show enhanced SE efficiency when treated with IAA or efflux-inhibited IAA, confirming that auxin efflux reduces the efficiency of Arabidopsis SE.
结果2-再生植株转化率：Importantly, in contrast to the 2,4-D system, where only 50–60% of the embryos converted to seedlings, all SEs induced by transport-inhibited natural auxins converted to seedlings.
结果3-不同再生系统：Efflux-inhibited IAA, like 2,4-D, also efficiently induced SE from carrot suspension cells, whereas IAA alone could not, and efflux-inhibited 4-Cl-IAA significantly improved de novo shoot regeneration in Brassica napus.
结论：Our data provides new insights into the action of 2,4-D as an efficient inducer of plant regeneration but also shows that replacing this synthetic auxin for efflux-inhibited natural auxin significantly improves different types of plant regeneration, leading to a more synchronized and homogenous development of the regenerated plants.
摘 要
植物基因组编辑和繁殖是作物育种和繁殖领域重要的工具。二者均严重依赖于高效的离体再生系统。在作物繁殖领域广泛应用的植物离体再生系统主要有两种，即体细胞胚胎发生体系和从头芽再生体系。在大多体胚发生或芽再生体系中，外植体通常需要经过高浓度的人工合成生长素类似物2,4-D的处理，而天然生长素IAA或者4-Cl-IAA的效果并不理想。基于前人的报道，相比于天然生长素，2,4-D不能有效的从植物细胞中被排出，因此作者在本文中测试了是否内源生长素的外排能够有助于诱导拟南芥未成熟胚胎进行体胚发生。作者发现，在施加生长素转运抑制剂NPA之后，无论是2,4-D还是植物天然生长素，均能够高效诱导拟南芥体胚发生。此外，pin2突变体或abcb1 abcb19双突的未成熟胚胎在IAA处理或者外排抑制的IAA处理，其体胚发生效率明显增强，表明生长素外排降低了拟南芥的体胚发生效率。重要的是，与2,4-D系统中仅50～60%的体细胞胚能够转化成再生植株不同，利用植物天然生长素辅以生长素转运抑制而获得的体细胞胚胎100%能够转化形成再生植株。2,4-D等外排抑制的IAA同样能够诱导胡萝卜悬浮细胞的体胚发生，而IAA单独则不能；另外，外排抑制的4-Cl-IAA能够显著提升油菜的从头芽再生能力。本文的研究揭示了人工合成生长素2,4-D作为植物再生的高效诱导剂的原因，同时揭示了利用外排抑制的自然生长素代替2,4-D可以改善不同类型的植物再生系统，从而使得再生植株的发育更加同步。
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