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Isolation and Genetic Mapping of a Non-Lethal Rice (Oryza sativa L.) low phytic acid 1 Mutation

^a USDA-ARS, Forage and Range Research Lab., Utah State Univ., Logan, UT 84322-6300 USA
^b USDA-ARS, Dale Bumpers National Rice Research Center, Stuttgart, AR 72610 USA
^c USDA-ARS, National Small Grains Germplasm Research Facility, Aberdeen, ID 83210 USA

View larger version (60K): [in a new window]   Fig. 1 Segregation of Orion M2-70 in the M2 and M3 generations. Seeds produced by a given plant were individually crushed and assayed for inorganic P. (A) M2 generation; 10 M3 seeds were tested for each of 10 Orion M2-70 M2 plants. (B) M3 generation; 10 M4 seeds were tested for each of 14 Orion M2-70 M3 plants. Standards: (1) 0.0 µg P; (2) 0.15 µg P; (3) 0.46 µg P; (4) 0.93 µg P; (5) 1.39 µg P. In this study, wild-type rice seeds typically have inorganic P levels ( = 0.15 mg PI g-1) that yield an assay result < Standard No. 2. Orion M2-70 seeds have inorganic P levels resulting in an assay = Standard No. 5, indicating these seeds contained > 1.5 mg Pi g-1

View larger version (59K): [in a new window]   Fig. 2 Segregation of Kaybonnet M2-2045 in the M2 through M4 generations. Seeds produced by a given plant were individually crushed, and assayed for inorganic P. (A) M2 generation; A single M3 seed from each of 10 plants comprising a given M2 row were crushed, and tested. The rows shown are M2-2041 through M2-2048. (B) M3 generation, M3-2045-1 through M3-2045-9; 10 M4 seeds from each of nine M3 plants were tested. (C) M4 generation; a total of eight M5 seeds were tested for each of five sibling M4 plants representing the nine M3s in (B). The first four M5 tests for each M4 are shown. Standards: (1) 0.0 µg P; (2) 0.15 µg P; (3) 0.46 µg P; (4) 0.93 µg P; (5) 1.39 µg P. In this study, wild-type rice seeds typically have inorganic P levels ( = 0.15 mg Pi g-1) that yield an assay result < Standard No. 2. Kaybonnet M2-2045 seeds have inorganic P levels that yield an assay result typically Standard No. 4, indicating these seeds contained 1.0 mg Pi g-1

View larger version (22K): [in a new window]   Fig. 3 Analysis of inositol phosphates in wild-type and lpa1-1rice seed. Standards or samples were loaded onto a strong-anion exchange column and eluted with a complex gradient. Ins Ps were detected following post-column derivitization via negative absorbance at 550 nm. (A) Elution of 100 nmol of myo-inositol 1,2,3,4,5,6-hexakisphosphate (Ins P6) standard. (B, C) Equal amounts of wild-type (B) or lpa1-1 (C) seed tissue were extracted, and equal aliquots of filtered supernatant were fractionated as in (A)

View larger version (31K): [in a new window]   Fig. 4 Diagramatic map of chromosomal regions containing the lpa1 and lpa2 loci and myo-inositol 1-phosphate synthase (MIPS) genes of barley, maize, and rice. As determined in this study, the map locations of rice lpa1-1 and MIPS gene are indicated on the rice chromosome 2 (outer right) and rice chromosome 3 (outer left) maps (Chen et al., 1997; Guiderdoni et al., 1992; Huang et al., 1994; Huang et al., 1997; and Panaud et al., 1997), respectively. Horizontal dashed lines indicate cross hybridization of RFLP probes, including MIPS (Ahn and Tanksley, 1993; Kurata et al., 1994; Larson and Raboy, 1999; VanDeynze et al., 1995) in addition to map locations of the maize and barley lpa1 and lpa2 mutations (Larson et al., 1998; Larson and Raboy, 1999)

View larger version (86K): [in a new window]   Fig. 5 A rice myo-inositol 1-phosphate synthase (MIPS) STS-PCR restriction polymorphism analyzed by 6% (w/v) PAGE. This polymorphism was mapped to rice chromosome 3 in the IR64 x Azucena doubled haploid mapping population