Journal Article

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

Li, Wei, Liu, Hongmei, Niu, Shihui, Ma, Jingjing, Guo, Yingtian, Liu, Shuangwei

2020

Overview

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 β-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 μM acetosyringone for 30 min followed by 3 days in the dark at 23 ± 1 °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.

• Publisher: BioMed Central,BioMed Central Ltd,Springer Nature B.V,BMC
• Subject: absorbance / Acetosyringone / Agrobacterium / Agrobacterium-mediated transformation / Analysis / Angiosperms / Arabidopsis / beta-glucuronidase / Biological Techniques / Biomedical and Life Sciences / coculture / Conifers / Cultivation / Efficiency / Efficient transformation system / Flowering / Flowers & plants / Gene expression / Genetic analysis / Genetic aspects / Genetic transformation / Genetics / Genomes / genomics / Gram-negative bacteria / GUS staining / Gymnosperms / heterologous gene expression / Identification and classification / Infections / Life Sciences / Methodology / Methods / Optical density / Pines / Pinus tabuliformis / Pinus tabuliformis callus / Plant bacterial diseases / Plant Sciences / Reporter gene / reporter genes / Seeds / staining / Statistical analysis / Transformations / transgenes / Transient gene expression / Trees / vegetative growth