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<p><i>Larix</i> is a valuable ecological and economic tree species that is essential to China’s timber supply and environmental conservation. However, genetic transformation of larch via <i>Agrobacterium Tumefaciens</i> has been severely limited by poor tissue culture responses and genotype dependency, making the process time-consuming and inefficient. <i>Agrobacterium rhizogenes</i>-mediated hairy root transformation presents a promising alternative for investigating the function of genes, particularly those root-specific genes. In this work, we describe a simple, reliable, and effective transformation approach using <i>A. rhizogenes</i> to generate composite larch plants for transgene expression analysis. Our findings demonstrate that <i>A. rhizogenes</i> enables stable generation of transgenic roots, which can maintain normal plant growth after removing non-transgenic roots. The results show that strain K599 is the most effective for hairy root transformation in larch. Using 50-day-old seedlings as explants and inoculating a 1.5&#xa0;cm slant cut of the residual hypocotyl at an optical density (OD<sub>600</sub>) of 1.5, we achieved a transformation efficiency of up to 25.19%. Furthermore, we conducted a comprehensive analysis of the ATP-binding cassette (ABC) transporter LkABCG36 to investigate its role in lignin synthesis using our transformation protocol. The results demonstrated that LkABCG36 enhances xylem formation in hairy roots by promoting the transport of lignin monomers. This technique offers a rapid and effective tool for in vivo gene function studies in larch.</p>

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Fluorescent Larix hairy root construction and its application to gene function in larch

  • 期刊论文

  • 发表日期：

    2025年4月8日

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  • Junying Lu,
  • Nan Sun,
  • Ming Tian,
  • Dadi Zhao,
  • Yuxiao Yu,
  • Xiangning Jiang,
  • Ying Gai

摘要

Larix is a valuable ecological and economic tree species that is essential to China’s timber supply and environmental conservation. However, genetic transformation of larch via Agrobacterium Tumefaciens has been severely limited by poor tissue culture responses and genotype dependency, making the process time-consuming and inefficient. Agrobacterium rhizogenes-mediated hairy root transformation presents a promising alternative for investigating the function of genes, particularly those root-specific genes. In this work, we describe a simple, reliable, and effective transformation approach using A. rhizogenes to generate composite larch plants for transgene expression analysis. Our findings demonstrate that A. rhizogenes enables stable generation of transgenic roots, which can maintain normal plant growth after removing non-transgenic roots. The results show that strain K599 is the most effective for hairy root transformation in larch. Using 50-day-old seedlings as explants and inoculating a 1.5 cm slant cut of the residual hypocotyl at an optical density (OD600) of 1.5, we achieved a transformation efficiency of up to 25.19%. Furthermore, we conducted a comprehensive analysis of the ATP-binding cassette (ABC) transporter LkABCG36 to investigate its role in lignin synthesis using our transformation protocol. The results demonstrated that LkABCG36 enhances xylem formation in hairy roots by promoting the transport of lignin monomers. This technique offers a rapid and effective tool for in vivo gene function studies in larch.