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Functional Analysis of a Phosphate Transporter Gene OsPtl in Rice

Author: HuangXinPeng
Tutor: SunShuBin;XuGuoHua
School: Nanjing Agricultural College
Course: Plant Nutrition
Keywords: rice OsPT1 yeast xenopus oocytes overexpression
CLC: S511
Type: Master's thesis
Year: 2010
Downloads: 10
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Abstract

Phosphorus is a constituent of key molecules such as ATP, nucleic acids, and phospholipids. It plays a crucial role in energy transfer, metabolic regulation, and protein activation. Although abundant phosphorus is present in many soils, phosphate (Pi) is one among the least-available macronutrients required by plants due to its low concentration in the soluble form and slow rate of diffusion to root surface. Enhancing expression of some phosphate transporter genes is one of the key mechanisms of rice to adapt the deficiency of phosphate. Understanding of the physiological functions of phosphate transporters is essential for dissecting the molecular mechanism of phosphate uptake, translocation and utilization.We observed the expression pattern of OsPT1 by using Real-Time PCR technique. We also obtained T-DNA inserted mutants of the gene from Korea and lines with overexpression of OsPT1. In addition, we provide electrophysiological evidence demonstrating the electrical activity in yeast and Xenopus oocytes heterologous expression systems. The main results are summarized as follows:Genomic DNA sequence analysis shows that OsPT1 do not have intron, OsPT1 is localized on chromosome 3 which has a single copy in the genome of Nipponbare. The deduced amino acid sequences of OsPTl suggest that OsPT1 is typical intergrating membrane protein and contains consensus site for N-glycosylation.Real-Time PCR indicates that the expression of OsPTl increased both in roots and leaves upon Pi starvation. The expression of OsPTl in leaves is higher than that in the roots which in dependent of Pi supplying.We tested the expression of OsPT1 proteins in yeast and Xenopus oocytes heterologous expression systems. OsPTl exhibits an apparent Km of 177μM and was able to complement Pi uptake mutant yeast in the high affinity concentration range. In oocyte expression system the Km is 27μM. OsPTl might be a hign Pi affinity phosphate transporter. The pH experiments in yeast and Xenopus oocytes heterologous expression systems suggest that the activity of OsPTl is dependent on electrochemical membrane potential mediates by H+-ATPase. The results indicated that OsPT1 might be a hign Pi affinity co-transporter of H+/Pi.OsPT1 overexpression enhanced absorption of phosphorus and increased biomass of the transgenics. OsPTl may be not only involved in the absorption of phosphorus but also involved in the transportation process.

Functional Analysis of a Phosphate Transporter Gene OsPtl in Rice

Abstract

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