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Influence of bacterial density during preculture on Agrobacterium-mediated transformation of tomato

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Abstract

An improved bacterial preculture protocol for Agrobacterium-mediated genetic transformation was developed for an economic tomato cultivar (Solanum lycopersicum L. cv. Zhongshu No. 4). Frequencies of transient gene expression and stable transformation were influenced by the density of Agrobacterium preculture and not the density of Agrobacterium used for infection. The improved protocol presented in this study depends on the use of an overnight-grown Agrobacterium preculture density of OD600 nm = 1.0, diluted 1/10th with Luria-Bertani (LB) liquid medium, and grown for an additional 4 h. Cultures are collected and resuspended in a liquid cocultivation medium-I, adjusted to OD600 nm = 0.1. Using this modified Agrobacterium preparation, transient β-glucuronidase expression was higher than 90%, and transformation efficiency reached 44.7%. This improved transformation is simple, repeatable, does not require a feeder layer, and most notably, the transformation frequency is stable and highly efficient.

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Acknowledgments

The authors thank Dr. Sumei Li from the Institute of Soil Science, Chinese Academy of Sciences, for valuable suggestions on transgenic techniques. The authors also thank the anonymous reviewers for their valuable comments and suggestions that had improved this manuscript greatly. This work was supported by the National Natural Science Foundation of China (40671100) and the National Basic Research Program of China (2007CB109303).

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Authors and Affiliations

  1. State Key Laboratory of Soil and Sustainable Agriculture, Institute of Soil Science, Chinese Academy of Sciences, 210008, Nanjing, China

    Nan Gao, Yu Cao, Yanhua Su & Weiming Shi

  2. Graduate School of the Chinese Academy of Sciences, 100039, Beijing, China

    Nan Gao & Yu Cao

  3. College of Chemistry and Environmental Science, Nanjing Normal University, 210097, Nanjing, China

    Weishou Shen

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  1. Nan Gao
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  2. Weishou Shen
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  3. Yu Cao
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  4. Yanhua Su
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Correspondence to Weiming Shi.

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Gao, N., Shen, W., Cao, Y. et al. Influence of bacterial density during preculture on Agrobacterium-mediated transformation of tomato. Plant Cell Tiss Organ Cult 98, 321–330 (2009). https://doi.org/10.1007/s11240-009-9566-2

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