Abstract
Mutant populations are indispensable genetic resources for functional genomics in all organisms. However, suitable rice mutant populations, induced either by chemicals or irradiation still have been rarely developed to date. To produce mutant pools and to launch a search system for rice gene mutations, we developed mutant populations of Oryza sativa japonica cv. Taichung 65, by treating single zygotic cells with N-methyl-N-nitrosourea (MNU). Mutagenesis in single zygotes can create mutations at a high frequency and rarely forms chimeric plants. A modified TILLING system using non-labeled primers and fast capillary gel electrophoresis was applied for high-throughput detection of single nucleotide substitution mutations. The mutation rate of an M2 mutant population was calculated as 7.4 × 10−6 per nucleotide representing one mutation in every 135 kb genome sequence. One can expect 7.4 single nucleotide substitution mutations in every 1 kb of gene region when using 1,000 M2 mutant lines. The mutations were very evenly distributed over the regions examined. These results indicate that our rice mutant population generated by MNU-mutagenesis could be a promising resource for identifying mutations in any gene of rice. The modified TILLING method also proved very efficient and convenient in screening the mutant population.
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Acknowledgments
The authors would like to thank Drs. BJ, Till, L. Comai and S, Henikoff for their kind introduction of the original tilling method for us. We also thank Drs. M. Yano, Y. Nagamura, and H. Hirochika for their support to this work, and Ms. K. Sasaga for assistance. The appreciations are extended to Drs. Y. Harushima and S. Yamaki for fruitful discussion, and also to Drs. E. Gilchrist (University of British Columbia), M. Fujita and Mr. Thirumurugan for critical reading of this manuscript and providing helpful comments. This work was supported by a grant from the Ministry of Agriculture, Forestry and Fisheries of Japan (Green technology Project QT1004), and the National Bio-resource Project from the Ministry of Education, Sport, Culture and Technology (MEXT), Japan.
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Communicated by Masahiro Yano.
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Suzuki, T., Eiguchi, M., Kumamaru, T. et al. MNU-induced mutant pools and high performance TILLING enable finding of any gene mutation in rice. Mol Genet Genomics 279, 213–223 (2008). https://doi.org/10.1007/s00438-007-0293-2
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DOI: https://doi.org/10.1007/s00438-007-0293-2