Molecular breeding for transgenic rice with low-phytic-acid phenotype through manipulating myo-inositol 3-phosphate synthase gene

In most plant seeds, phosphorus is stored primarily as myo -inositol 1,2,3,4,5,6-hexa kis phosphate (InsP 6 ; phytic acid). Reducing the phytic acid content of seeds is a major breeding target, both to increase the availability of mineral nutrients and to decrease the environmental load of phosphorus. The first step in phytic acid biosynthesis and inositol metabolism is catalyzed by 1 d - myo -inositol 3-phosphate (Ins(3)P 1 ) synthase. In this study, we aimed to reduce phytic acid levels in rice seeds by manipulating the expression of the rice Ins(3)P 1 synthase gene RINO1 using transgenic methods. RINO1 cDNA was transformed into rice plants in the antisense orientation under the control of the rice major storage protein glutelin GluB-1 promoter. The T 4 generation of a stable transgenic line that contained four copies of the transgene showed little morphological differences compared to non-transgenic rice. In the T 5 seeds of this line, severe reductions in RINO1 protein levels were observed during the late maturing stages of ripening. Most of the T 5 seeds contained higher amounts of inorganic phosphates (Pi), without a reduction in total phosphorus levels, compared to non-transgenic seeds. Ion chromatography analysis suggested that the increase in available Pi is accompanied by a molar-equivalent decrease in phytic acid P. The expression patterns of RINO1 and GluB-1 were examined by quantitative real-time reverse transcriptase-polymerase chain-reaction (RT-PCR). Potential strategies for further molecular breeding to reduce phytic acid levels in seeds are discussed.