The Study Of Overexpression Of OsPHF1to Improve Phosphate Uptake Ability In Rice(Orrvza Sativa L.)

The phosphate (Pi) absorbed by plants is very limited because of the relative inmobility of phosphorus and the low level of Pi in soils. Rice is one of the most important cereal crops in the world. The lower utilization of P-fertilizer is one of main factors limiting rice growth and production. Therefore, breeding Pi-efficient crops including rice is importance of sustainable agriculture. Previous studies have shown that PHF1plays an important role in the trafficking of Pi transporters from the endoplasmic reticulum (ER) to plasma membrane (PM), which influences Pi uptake and translocation in rice. It also indicates that overexpression of OsPHFl can increase Pi uptake ability in rice.In this study, we developed transgenic plants of9311, an indica restorer line of Super Hybrid Rice, with overexpression of OsPHFl using glyphosate resistant gene, by Agrobacterium-mediated rice transformation. Then we used a nested integrated PCR method to identify the T-DNA insertion site of the transgenic lines. Flanking sequencing analysis revealed two lines with single insertional site of transformed OsPHFl. The two lines were used for further evaluation for Pi-uptake efficiency and tolerance to lower P stress. The main results were as follows:1. The flanking sequence analysis indicates that the insertional OsPHFl is integrated into the34404kbp of chromosome4and the2736kbp of chromosome6, respectively. To determine the single insertional site, we tested the segregation of the insertional site using the self-fertilized progenies from TO generation. The results showed that the segregations of both lines fit Mendelian segregation, indicating the single copy lines.2. Field test for T1generation of the two transgenic plants (designated as9311/PHF1(O)-1and9311/PHF1(O)-3) showed significant higher yield, biomass and panicle number compared with the wild type in low P soil (Olsen-Pi:3.5ppm, soil pH:7.1). The yield of the transgenic plants increased by about25%in the low P soil. There’s no significant difference between the transgenic plants and the wide type in the high P soil (Olsen-Pi:9.4ppm, soil pH:6.0).3. In Pi-supplied solution cultures (200μM), we analysised the T2generation homozygous lines and widetype9311by RT-PCR, qRT-PCR, Western blot using anti-PHF1antibody and Pi concentrations. The results showed that the mRNA and protein levels of OsPHFl in the transgenic lines were significantly increased and concomitantly Pi-uptake ability is increased compared with the wild type plants.In conclusion, our results indicate that overexpressed OsPHFl in9311can improve Pi upake ability.