An efficient system for Agrobacterium-mediated transient transformation in Pinus tabuliformis.

Background: Functional genomic studies using genetics approaches of conifers are hampered by the complex and enormous genome, long vegetative growth period, and exertion in genetic transformation. Thus, the research carried out on gene function in Pinus tabuliformis is typically performed by heterologous expression based on the model plant Arabidopsis. However, due to the evolutionary and vast diversification from non-flowering (gymnosperms) to flowering (angiosperms) plants, several key differences may alter the underlying genetic concerns and the analysis of variants. Therefore, it is essential to develop an efficient genetic transformation and gene function identification protocol for P. tabuliformis. Results: In the present study we established a highly efficient transgene Agrobacterium-mediated transient expression system for P. tabuliformis. Using a beta-glucuronidase gene (GUS) as a reporter gene expression, the highest transformation efficiency (70.1%) was obtained by co-cultivation with Agrobacterium strain GV3101 at an optical density at 600 nm of 0.8, with 150 muM acetosyringone for 30 min followed by 3 days in the dark at 23 +/- 1 degrees C. This protocol would be applied to other conifers; GUS staining was observed 24 h post-infection. Conclusions: We report a simple, fast, and resilient system for transient Agrobacterium-mediated transformation high-level expression of target genes in P. tabuliformis, which will also improve transformation efficiency in other conifer species.