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Identification and characterization of the soybean IPK1 ortholog of a low phytic acid mutant reveals an exon-excluding splice-site mutation

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Abstract

Phytic acid (myo-inositol 1, 2, 3, 4, 5, 6 hexakisphosphate) is an important constituent of soybean meal. Since phytic acid and its mineral salts (phytates) are almost indigestible for monogastrics, their abundance in grain food/feed causes nutritional and environmental problems; interest in breeding low phytic acid has therefore increased considerably. Based on gene mapping and the characteristics of inositol polyphosphates profile in the seeds of a soybean mutant line Gm-lpa-ZC-2, the soybean ortholog of inositol 1,3,4,5,6 pentakisphosphate (InsP5) 2-kinase (IPK1), which transforms InsP5 into phytic acid, was first hypothesized as the candidate gene responsible for the low phytic acid alteration in Gm-lpa-ZC-2. One IPK1 ortholog (Glyma14g07880, GmIPK1) was then identified in the mapped region on chromosome 14. Sequencing revealed a G → A point mutation in the genomic DNA sequence and the exclusion of the entire fifth exon in the cDNA sequence of GmIPK1 in Gm-lpa-ZC-2 compared with its wild-type progenitor Zhechun No. 3. The excluded exon encodes 37 amino acids that spread across two conserved IPK1 motifs. Furthermore, complete co-segregation of low phytic acid phenotype with the G → A mutation was observed in the F2 population of ZC-lpa x Zhexiandou No. 4 (a wild-type cultivar). Put together, the G → A point mutation affected the pre-mRNA splicing and resulted in the exclusion of the fifth exon of GmIPK1 which is expected to disrupt the GmIPK1 functionality, leading to low phytic acid level in Gm-lpa-ZC-2. Gm-lpa-ZC-2, would be a good germplasm source in low phytic acid soybean breeding.

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Acknowledgments

We thank Dr. Hairui Cui for his suggestions regarding the design of RT-PCR experiments; we are also grateful to Prof. Pengyin Chen for his comments and suggestions on the early versions of this manuscript. The research was financially supported by the China Natural Science Foundation through research contract No. 30871542 to FJY and in part supported by the Fundamental Research Funds for Central Universities, the Special Fund for Agro-scientific Research in the Public Interest (201103007) and by the Sino-Swiss Joint Research Project (2009 DFA32040) to QYS. Our heartfelt thanks go to the anonymous reviewers who offered their critical comments for the improvement of this paper.

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Authors and Affiliations

  1. Institute of Crop Science and Nuclear Technology Utilization, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, China

    Feng-Jie Yuan, Dan-Hua Zhu, De-Kun Dong, Xu-Jun Fu, Shen-Long Zhu & Bai-Quan Li

  2. Key Laboratory of Nuclear Agricultural Sciences of Ministry of Agriculture and Zhejiang Province, and IAEA-Zhejiang University Collaborating Center, Institute of Nuclear Agricultural Sciences, Zhejiang University, Hangzhou, 310029, China

    Yuan-Yuan Tan & Qing-Yao Shu

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  1. Feng-Jie Yuan
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  2. Dan-Hua Zhu
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Correspondence to Qing-Yao Shu.

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Yuan, FJ., Zhu, DH., Tan, YY. et al. Identification and characterization of the soybean IPK1 ortholog of a low phytic acid mutant reveals an exon-excluding splice-site mutation. Theor Appl Genet 125, 1413–1423 (2012). https://doi.org/10.1007/s00122-012-1922-7

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