MNU-induced mutant pools and high performance TILLING enable finding of any gene mutation in rice

Mol Genet Genomics. 2008 Mar;279(3):213-23. doi: 10.1007/s00438-007-0293-2. Epub 2007 Oct 19.

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 M(2) mutant population was calculated as 7.4 x 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 M(2) 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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Base Sequence
  • DNA Mutational Analysis / methods
  • DNA Primers / genetics
  • DNA, Plant / genetics
  • Genes, Plant / drug effects
  • Genetic Techniques*
  • Methylnitrosourea / toxicity
  • Mutagenesis
  • Mutagens / toxicity
  • Mutation*
  • Oryza / drug effects
  • Oryza / genetics*
  • Polymerase Chain Reaction

Substances

  • DNA Primers
  • DNA, Plant
  • Mutagens
  • Methylnitrosourea