PJB-2024-105
A Simple, Speedy and Straightforward Method for the Genetic Transformation of Poplar with Agrobacterium rhizogenes
Mei Han
Abstract
Poplars are one of the most widespread ligneous plants in the world, which have a variety of applications ranging from landscape, agriculture, and industry to research. Although Agrobacterium tumefaciens-mediated transformation method has been widely utilized for poplar genetic manipulation, many technique defects such as the time-consuming procedures, and the occurrence of chimeric transformant have emerged. To address this problem, we developed a simple, fast, straightforward and effective gene transformation method by using Agrobacterium rhizogenes to induce hairy roots in poplars, and obtained regenerated-transgenic poplar plantlets within two to three months. The optimal procedure involved the usage of poplar apex buds as explants, infection of A. rhizogenes by directly pricking colonies with the cuttings on the agar plates, and improving hairy-root initiation by application of indole-3-butyric acid (IBA) in combination with hairy-root morphological identification. Moreover, the GFP marker gene was employed to track record the expression level of the introduced genes using a portable fluorescence lamp. Further quantitative real-time PCR and the enzyme activity assay showed that the transcript abundance and enzyme activity of our target gene (i.e., aspartate aminotransferase) were significantly elevated in the transgenic plants regenerated from the hairy roots when compared with the non-transformed control, confirming that the current A. rhizogenes-mediated poplar transformation method is effective and applicable for poplar genetic manipulation and functional genomic study.
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