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A rice gene activation/knockout mutant resource for high throughput functional genomics

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Abstract

Using transfer DNA (T-DNA) with functions of gene trap and gene knockout and activation tagging, a mutant population containing 55,000 lines was generated. Approximately 81% of this population carries 1–2 T-DNA copies per line, and the retrotransposon Tos17 was mostly inactive in this population during tissue culture. A total of 11,992 flanking sequence tags (FSTs) have been obtained and assigned to the rice genome. T-DNA was preferentially (∼80%) integrated into genic regions. A total of 19,000 FSTs pooled from this and another T-DNA tagged population were analyzed and compared with 18,000 FSTs from a Tos17 tagged population. There was difference in preference for integrations into genic, coding, and flanking regions, as well as repetitive sequences and centromeric regions, between T-DNA and Tos17; however, T-DNA integration was more evenly distributed in the rice genome than Tos17. Our T-DNA contains an enhancer octamer next to the left border, expression of genes within genetics distances of 12.5 kb was enhanced. For example, the normal height of a severe dwarf mutant, with its gibberellin 2-oxidase (GA2ox) gene being activated by T-DNA, was restored upon GA treatment, indicating GA2ox was one of the key enzymes regulating the endogenous level of GA. Our T-DNA also contains a promoterless GUS gene next to the right border. GUS activity screening facilitated identification of genes responsive to various stresses and those regulated temporally and spatially in large scale with high frequency. Our mutant population offers a highly valuable resource for high throughput rice functional analyses using both forward and reverse genetic approaches.

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Acknowledgments

We thank Drs Tuan-Hua David Ho, Anthony H.C. Huang, and Kenrick Deen for their critical review of this manuscript and Drs Hua-Mei Chang and Peng-Wen Chen and Mr Cheng-Tung Lo from Vita Genomics, and Ms Lin-Chih Yu for technical assistance. This work was supported by grants from Academia Sinica (AS91IB2PP, AS90-AB-IMB-01, AS91-AB-IMB-01, AS92-AB-IMB-03, and AS93-AB-IMB-02 to Su-May Yu and 92S510921 and 92S508932-G to Yue-Ie Hsing) and the National Science Council (NSC92-2317-B001-032 and NSC92-2317-B-001-036 to Su-May Yu) of the Republic of China.

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Authors and Affiliations

  1. Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei, 115, Taiwan, ROC

    Yue-Ie Hsing, Po-Chang Lu, Chun-I Chung, Yao-Cheng Lin & Ai-Ling Hour

  2. Taiwan Agricultural Research Institute, Wufeng, Taichung, 413, Taiwan, ROC

    Chyr-Guan Chern, Shu Chen, Hsing-Mu Yen, Charng-Pei Li, Chiu-Kai Wey, Ching-Shan Tseng, Ming-Hsing Lai & Sheng-Chung Huang

  3. Department of Biotechnology and Bioinformatics, Asia University, Lioufeng Road, Wufeng, Taichung , 413, Taiwan, ROC

    Ming-Jen Fan

  4. Institute of Molecular Biology, Academia Sinica, Nankang, Taipei , 115, Taiwan, ROC

    Ku-Ting Chen, Shuen-Fang Lo, Peng-Kai Sun, Kuo-Wei Lee, Yi-Chieh Wang, Jyh-Long Chen, Yet-Walt Wang, Ya-Chi Chang, Min-Wei Tsai, Yi-Show Lin, Yin-Chin Chen & Su-May Yu

  5. Institute of Molecular Biology, National Chung-Hsing University, Wufeng, Taichung , 400, Taiwan, ROC

    Shuen-Fang Lo & Liang-Jwu Chen

  6. Department of Agronomy, National Chia-Yi University, Chia-Yi, 600, Taiwan, ROC

    Shin-Lon Ho

  7. Department of Biotechnology, Fooyin University, Kaohsiung Hsien , 831, Taiwan, ROC

    Wen-Lii Huang

  8. Academia Sinica Biotechnology Experimental Center in Southern Taiwan, Southern Taiwan Science Park, Tainan , 741, Taiwan, ROC

    Swee-Suak Ko

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  1. Yue-Ie Hsing
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Correspondence to Su-May Yu.

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Yue-Ie Hsing, Chyr-Guan Chern, and Ming-Jen Fan have contributed equally.

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Hsing, YI., Chern, CG., Fan, MJ. et al. A rice gene activation/knockout mutant resource for high throughput functional genomics. Plant Mol Biol 63, 351–364 (2007). https://doi.org/10.1007/s11103-006-9093-z

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