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Identification and characterization of a novel Waxy allele from a Yunnan rice landrace

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Abstract

Low amylose content (AC) is a desirable trait for rice (Oryza sativa L.) cooking quality and is selected in soft rice breeding. To gain a better understanding of the molecular mechanism controlling AC formation, we screened 83 Yunnan rice landraces in China and identified a rice variety, Haopi, with low AC. Genetic analyses and transgenic experiments revealed that low AC in Haopi was controlled by a novel allele of the Wx locus, Wx hp, encoding a granule-bound starch synthase (GBSSI). Sequence comparisons of Wx hp and Wx b alleles (from Nipponbare) showed several nucleotide changes in the upstream regulatory regions (including the promoter, 5′-untranslated region, and first intron 5′ splicing junction site). Interestingly, these changes had no obvious effect on the expression level and splicing efficiency of Wx transcripts. In addition, an examination of the coding region revealed that the Wx hp allele carries an A-to-G change at nucleotide position +497 from the start codon, resulting in an Asp165/Gly165 substitution. The amino acid substitution had no detectable effects on GBSSI activity in vitro; however, it notably reduced the binding of GBSSI to starch granules, resulting in a reduction of AC in rice seeds. Moreover, three other Yunnan landraces with low AC also carry a nucleotide substitution identical to Haopi at the +497 position of the Wx gene, suggesting common ancestry. Based on the single-nucleotide polymorphism, we have developed a new derived cleaved amplified polymorphic sequence marker for use in breeding practice to manipulate AC in rice endosperm.

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Abbreviations

AC:

Amylose content

Wx:

Waxy

GBSSI:

Granule-bound starch synthase I

SNP:

Single nucleotide polymorphism

dCAPS:

Derived cleaved amplified polymorphic sequence

3D:

Three dimension

RT-PCR:

Reverse transcriptase polymerase chain reaction

DAF:

Days after flowering

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Acknowledgments

We thank Dr. Haiyang Wang (Boyce Thompson Institute for Plant Research, Cornell University) and Dr. Tianbao Yang (Center for Integrated Biotechnology and Department of Horticulture, Washington State University) for proofreading and commenting on the manuscript. We also thank Dr. Mao Long (Institute of Crop Science, Chinese Academy of Agricultural Sciences, China) for his helpful comments on the manuscript and starch synthase enzymatic assays. This work was supported by grants from the National High Technology Research and Development Program of China (No. 2006AA100101, 2006AA10Z1A5, 2007AA10Z116, 2006BAD01A01-5), the National Natural Science Foundation of China (No. 30270811 and 30700497), and the 111 project (B08025).

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Authors and Affiliations

  1. State Key Laboratory for Crop Genetics & Germplasm Enhancement, Jiangsu Provincial Center of Plant Gene Engineering, Nanjing Agricultural University, Weigang 1, 210095, Nanjing, China

    Linglong Liu, Xiaodong Ma, Shijia Liu, Changlan Zhu, Ling Jiang, Yihua Wang, Yi Shen, Yulong Ren, Hui Dong, Liangming Chen, Xi Liu, Zhigang Zhao & Jianmin Wan

  2. Institute of Crop Science, The National Key Facility for Crop Gene Resources and Genetic Improvement, Chinese Academy of Agricultural Sciences, 10081, Beijing, China

    Huqu Zhai & Jianmin Wan

Authors
  1. Linglong Liu
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  2. Xiaodong Ma
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  3. Shijia Liu
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  4. Changlan Zhu
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  5. Ling Jiang
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  9. Hui Dong
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  10. Liangming Chen
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  11. Xi Liu
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  12. Zhigang Zhao
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  13. Huqu Zhai
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  14. Jianmin Wan
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Corresponding author

Correspondence to Jianmin Wan.

Additional information

Linglong Liu and Xiaodong Ma have contributed equally to this work.

Accession Numbers

The GenBank accession number for the full-length Wx hp gene is EU871049.

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Liu, L., Ma, X., Liu, S. et al. Identification and characterization of a novel Waxy allele from a Yunnan rice landrace. Plant Mol Biol 71, 609–626 (2009). https://doi.org/10.1007/s11103-009-9544-4

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  • DOI: https://doi.org/10.1007/s11103-009-9544-4

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