SummarySoybean [Glycine max (L.) Merr.] is an exceptionally rich in isoflavones, and these compounds attach to oestrogen receptors in the human body, lessening the risk of breast cancer and effectively alleviating menopausal syndrome symptoms. Uncovering the molecular mechanisms that regulate soybean isoflavone accumulation is crucial for enhancing the production of these compounds. In this study, we combined bulk segregant analysis sequencing (BSA‐seq) and a genome‐wide association study (GWAS) to discover a novel R2R3‐MYB family gene, GmMYB77, that regulates isoflavone accumulation in soybean. Using the soybean hairy root transient expression system, we verified that GmMYB77 inhibits isoflavone accumulation. Furthermore, knocking out GmMYB77 significantly increased total isoflavone (TIF) content, particularly malonylglycitin, while its overexpression resulted in a notable decrease in contents of malonylglycitin and TIF. We found that GmMYB77 can directly binds the core sequence GGT and suppresses the expression of the key isoflavone biosynthesis genes Isoflavone synthase 1 (GmIFS1), Isoflavone synthase 2 (GmIFS2), Chalcone synthase 7 (GmCHS7) and Chalcone synthase 8 (GmCHS8) by using dual‐luciferase assays, electrophoretic mobility shift assays and yeast one‐hybrid experiments. Natural variations in the promoter region of GmMYB77 affect its expression, thereby regulating the malonylglycitin and TIF contents. Hap‐P2, an elite haplotype, plays a pivotal role in soybean breeding for substantially enhanced isoflavone content. These findings enhance our understanding of the genes influencing soybean isoflavone content and provide a valuable genetic resource for molecular breeding efforts in the future.

中文翻译：

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R2R3 型 MYB 转录因子 GmMYB77 负向调节大豆中异黄酮的积累 [Glycine max （L.） Merr.]


摘要大豆[Glycine max （L.） Merr.]是一种特别丰富的异黄酮，这些化合物附着在人体内的雌激素受体上，降低患乳腺癌的风险，有效缓解更年期综合症症状。揭示调节大豆异黄酮积累的分子机制对于增强这些化合物的产生至关重要。在这项研究中，我们将大量分离分析测序 （BSA-seq） 和全基因组关联研究 （GWAS） 相结合，发现了一种新的 R2R3-MYB 家族基因 GmMYB77，它调节大豆中异黄酮的积累。使用大豆毛状根瞬时表达系统，我们验证了 GmMYB77 抑制异黄酮积累。此外，敲除 GmMYB77 可显著提高总异黄酮 （TIF） 含量，尤其是丙二酰甘氨酸，而其过表达导致丙二酰甘氨酸和 TIF 含量显著降低。我们发现 GmMYB77 可以直接结合核心序列 GGT，并通过使用双荧光素酶测定、电泳迁移率变化测定和酵母单杂交实验抑制关键异黄酮生物合成基因异黄酮合酶 1 （GmIFS1）、异黄酮合酶 2 （GmIFS2）、查尔酮合酶 7 （GmCHS7） 和查尔酮合酶 8 （GmCHS8） 的表达。GmMYB77 启动子区的自然变化会影响其表达，从而调节丙二酰甘氨酸和 TIF 含量。Hap-P2 是一种优良的单倍型，在大豆育种中起着关键作用，可显著提高异黄酮含量。这些发现增强了我们对影响大豆异黄酮含量的基因的理解，并为未来的分子育种工作提供了宝贵的遗传资源。