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Generation of Rice Mutants by Chemical Mutagenesis

  • Thomas H. TaiEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 956)

Abstract

Chemical mutagenesis of rice has been used extensively to generate useful genetic variation for the purpose of breeding improved varieties. More recently, advances in high-throughput genotyping platforms have enabled the efficient detection of point mutations generated by chemical agents. This in turn has renewed interest in using traditional chemical mutagenesis to generate mutant populations for gene discovery and functional characterization. Targeting of Induced Local Lesions in Genomes (TILLING) is a powerful reverse genetics method which combines chemical mutagenesis with the high-throughput discovery of point mutations. Numerous chemical mutagens have been shown to be effective in generating point mutations and small deletions in rice. This chapter describes the use of a combination of sodium azide (NaN3) and N-nitroso-N-methylurea to generate populations that are suitable for TILLING as well as forward genetics and mutation breeding.

Key words

Chemical mutagenesis Reverse genetics Forward genetics TILLING Mutation breeding 

Notes

Acknowledgements

This protocol was developed with the technical assistance of P.M. Colowit and supported by the USDA Agricultural Research Service CRIS Project 5306-21000-016/17-00D and grant 2004-353604-14265 from the USDA Cooperative State Research, Education, and Extension Service, NRI Plant Genome Program.

References

  1. 1.
    Rutger JN (1983) Applications of induced and spontaneous mutation in rice breeding and genetics. In: Brady NC (ed) Advances in agronomy. Academic, New York, NY, pp 383–413Google Scholar
  2. 2.
    Tai TH (2007) Induced mutation in rice (Oryza sativa L.). Israel J Plant Sci 55:137–145CrossRefGoogle Scholar
  3. 3.
    Henikoff S, Comai L (2003) Single-nucleotide mutations for plant functional genomics. Annu Rev Plant Biol 54:37–401CrossRefGoogle Scholar
  4. 4.
    Zhang Q, Li J, Xue Y, Han B, Deng XW (2008) Rice 202: a call for an international coordinated effort in rice functional genomics. Mol Plant 1:715–719PubMedCrossRefGoogle Scholar
  5. 5.
    Henikoff S, Till BJ, Comai L (2004) TILLING. Traditional mutagenesis meets functional genomics. Plant Physiol 135:630–636PubMedCrossRefGoogle Scholar
  6. 6.
    Rigola D, van Oeveren J, Janssen A, Bonne A, Schneiders H, van der Poel HJA, van Orsouw NJ, Hogers RCJ, de Both MTJ, van Eijk MJT (2009) High-throughput detection of induced mutations and natural variation using KeyPoint™ technology. PLoS One 4:e4761PubMedCrossRefGoogle Scholar
  7. 7.
    Koornneef M (2002) Classical mutagenesis in higher plants. In: Gilmartin PM, Bowler C (eds) Molecular plant biology: a practical approach, vol 1. Oxford University, Oxford, UK, pp 1–11Google Scholar
  8. 8.
    Li SL, Redei GP (1969) Estimation of mutation rate in autogamous diploids. Radiat Bot 9:125–131CrossRefGoogle Scholar
  9. 9.
    Yamaguchi H (1962) The chimaeric formation in an X1 panicle after irradiation of dormant rice seed. Radiat Bot 2:71–77CrossRefGoogle Scholar
  10. 10.
    Leyser O (2000) Mutagenesis. In: Tucker GA, Roberts JA (eds) Methods in molecular biology: plant hormone protocols. Humana, Totowa, NJ, pp 133–144CrossRefGoogle Scholar
  11. 11.
    Kim Y, Schumaker KS, Zhu J (2006) EMS mutagenesis of Arabidopsis. In: Salinas J, Sanchez-Serrano JJ (eds) Methods in Molecular Biology: Arabidopsis Protocols, 2nd edn. Humana, Totowa, NJ, pp 101–103CrossRefGoogle Scholar
  12. 12.
    Maple J, Møller SG (2007) Mutagenesis in Arabidopsis. In: Rosato E (ed) Methods in Molecular Biology: Circadian Rhythmns: Methods and Protocols. Humana, Totowa, NJ, pp 197–206Google Scholar
  13. 13.
    Till BJ, Cooper J, Tai TH, Colowit P, Greene EA, Henikoff S, Comai L (2007) Discovery of chemically induced mutations in rice by TILLING. BMC Plant Biol 7:19PubMedCrossRefGoogle Scholar
  14. 14.
    Wu JL, Wu C, Lei C, Baraoidan M, Bordeos A, Madamba MR, Ramos-Pamplona M, Mauleon R, Portugal A, Ulat VJ, Bruskiewich R, Wang G, Jan Leach J, Khush G, Leung H (2005) Chemical-and irradiation-induced mutants of indica rice IR64 for forward and reverse genetics. Plant Mol Biol 59:85–97PubMedCrossRefGoogle Scholar
  15. 15.
    Awan MA, Konzak CF, Rutger JN, Nilan RA (1980) Mutagenic effects of sodium azide in rice. Crop Sci 20:663–668CrossRefGoogle Scholar
  16. 16.
    Suzuki T, Eiguchi M, Kumamaru T, Satoh H, Matsusaka H, Moriguchi K, Nagato Y, Kurata N (2008) MNU-induced mutant pools and high performance TILLING enable finding any gene mutation in rice. Mol Genet Genomics 279:213–223PubMedCrossRefGoogle Scholar
  17. 17.
    Owais WM, Kleinhofs A (1988) Metabolic activation of the mutagen azide in biological systems. Mutat Res 197:31–323Google Scholar
  18. 18.
    Olsen O, Wang X, von Wettstein D (1993) Sodium azide mutagenesis: preferential generation of A·T  →  G·C transitions in the barley Ant18 gene. Proc Natl Acad Sci USA 90:8043–8047PubMedCrossRefGoogle Scholar
  19. 19.
    Talame V, Bovina R, Sanguineti MC, Tuberosa R, Lundqvist U, Salvi S (2008) TILLMore, a resource for the discovery of chemically induced mutants in barley. Plant Biotech J 6:477–485CrossRefGoogle Scholar
  20. 20.
    Suzuki T, Moriguchi K, Tsuda K, Eiguchi M, Kumamaru T, Satoh H, Kurata N (2010) Neighboring nucleotide bias around MNU-induced mutations in rice. Rice Genet Newsl 25:90–91Google Scholar
  21. 21.
    Szarejko I, Maluszynski M (1999) High frequency of mutations after mutagenic treatment of barley seeds with NaN3 and MNH with application of inter-incubation germination period. Mutat Breed Newsl 44:28–30Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2013

Authors and Affiliations

  1. 1.USDA-ARS Crops Pathology and Genetics Research Unit and Department of Plant SciencesUniversity of CaliforniaDavisUSA

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