Department of Agronomy and Plant Breeding Sciences, University of Tehran, College of Aburaihan, Tehran, Iran; Centre for Integrative Legume Research, School of Agriculture and Food Sciences, The University of Queensland, Brisbane, Australia.
Centre for Integrative Legume Research, School of Agriculture and Food Sciences, The University of Queensland, Brisbane, Australia.
J Plant Physiol. 2021 Dec;267:153543. doi: 10.1016/j.jplph.2021.153543. Epub 2021 Oct 14.
Autoregulation of nodulation (AON) plays a central role in nodulation by inhibiting the formation of excess number of legume root nodules. In this study, the effect of hydroxymethylglutaryl-coenzyme A reductase 1 (GmHMGR1) gene expression on nodulation and the AON system in Glycine max (L.) Merr was investigated. Wild-type soybean (cultivar Bragg) and its near-isogenic supernodulating mutant (nitrate tolerant symbiotic) nts1007 were selected to identify the expression pattern of this gene in rootlets after inoculation by its microsymbiont Bradyrhizobium. For further analysis, the full length of GmHMGR1 and its promoter were cloned after amplification by inverse-PCR and BAC library screening. Also, we constructed an intron hairpin RNA interference (ihpRNAi) and a GmHMGR1 promoter: β-glucuronidase fusion constructs, consequently for suppression of GmHMGR1 and histochemical analysis in transgenic soybean hairy roots induced by Agrobacterium rhizogenes strain K599. The GmHMGR1 gene was functional during the early stages of nodulation with the AON system having a negative effect on GmHMGR1 expression and nodule formation in wild-type rootlets. GmHMGR1 was particularly expressed in the developing phloem within the root, nodules and nodule lenticels. Expression of GmHMGR1 in transgenic hairy roots was suppressed by RNAi silencing approximately 85% as compared to empty vector controls. This suggests that the GmHMGR1 gene has an important role in triggering nodule formation as its suppression caused a reduction of nodule formation in nts mutant lines with a deficient AON system.
自身调节结瘤作用（AON）在结瘤中起着核心作用，可抑制形成过多的豆科植物根瘤。本研究调查了羟甲基戊二酰基辅酶 A 还原酶 1（GmHMGR1）基因表达对大豆（Glycine max（L.）Merr）结瘤和 AON 系统的影响。选择野生型大豆（品种 Bragg）及其近等基因超结瘤突变体（耐硝酸盐共生）nts1007，以鉴定其共生根瘤菌接种后根毛中该基因的表达模式。为了进一步分析，通过反转 PCR 和 BAC 文库筛选扩增后克隆了全长 GmHMGR1 及其启动子。此外，我们构建了内含子发夹 RNA 干扰（ihpRNAi）和 GmHMGR1 启动子：β-葡萄糖醛酸酶融合构建体，用于抑制 GmHMGR1，并在根瘤农杆菌 K599 诱导的转基因大豆毛状根中进行组织化学分析。在 AON 系统的负调控下，GmHMGR1 基因在结瘤的早期阶段发挥功能，对野生型根毛中的 GmHMGR1 表达和结瘤形成有影响。GmHMGR1 在发育中的韧皮部、根、根瘤和根瘤皮孔中特异性表达。与空载体对照相比，RNAi 沉默使转基因毛状根中的 GmHMGR1 表达抑制约 85%。这表明 GmHMGR1 基因在触发结瘤形成中具有重要作用，因为其抑制导致 AON 系统缺陷的 nts 突变体系中结瘤形成减少。
