Construction of Yeast Two-hybrid cDNA Library and Screening of Interacting Proteins of SOC1a in Soybean Shoot Apices and Axillary Buds

doi:10.11983/CBB25062

Abstract

Abstract: INTRODUCTION: The shoot apices and axillary buds determine crop growth and yield potential, with their developmental states directly shaping shoot architecture. However, there are currently few cDNA libraries constructed for shoot apices and/or axillary buds in soybean ( Glycine max). RATIONALE: By constructing cDNA libraries for shoot apices and axillary buds, we can gain an in-depth understanding of the core mechanisms underlying plant architecture in soybean at the molecular level, thereby providing theoretical foundations and genetic resources for the design of high-yielding and well-adapted soybean varieties. RESULTS: This study constructed a yeast two-hybrid (Y2H) nuclear system cDNA library using shoot apices and axillary buds from the cultivar Williams 82 grown under long-day and short-day conditions at different developmental stages. Equal amounts of RNA extracted from these tissues were pooled and subjected to cDNA library construction using the Gateway method, followed by transcript diversity analysis. The resultant library had a capacity of 1.2×10⁷ CFU, with 100% recombination rate and an average length exceeding 1 000 bp of the inserted fragments, covering 29 170 genes. This cDNA library meets the library construction standards and is suitable for subsequent Y2H screening. Using the key floral transition and shoot architecture regulator SOC1a as a bait, we first tested the toxicity and self-activation of the recombinant pGBKT7-SOC1a and then performed library screening. A total of 32 positive clones were obtained, and after DNA sequencing, BLAST alignment, and functional annotation, 14 candidate interacting proteins were identified. Among them, five encoded genes were cloned into pGADT7 vector and subjected to pairwise retransformation assays with pGBKT7-SOC1a, confirming physical interactions between two of these proteins and SOC1a. Furthermore, the interaction between SOC1a and one of the candidate proteins, SEP2, was demonstrated through co-immunoprecipitation and luciferase complementation imaging assays. CONCLUSION: This study establishes a high-quality Y2H cDNA library for soybean meristematic tissues and identifies novel SOC1a-interacting proteins, providing critical molecular insights into SOC1a-mediated regulation of soybean shoot architecture development.

Key words: soybean, shoot apices and axillary buds, cDNA library, SOC1a, yeast two-hybrid

Huan Huang, Jiali Zhang, Xue Yang, Liyu Chen, Lin Yue, Baohui Liu, Hui Yang. Construction of Yeast Two-hybrid cDNA Library and Screening of Interacting Proteins of SOC1a in Soybean Shoot Apices and Axillary Buds[J]. Chinese Bulletin of Botany, 2026, 61(3): 386-401.

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URL: https://www.chinbullbotany.com/EN/10.11983/CBB25062

https://www.chinbullbotany.com/EN/Y2026/V61/I3/386

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