新磷酸转运蛋白基因EcPT1的克隆与鉴定

Y. Zhong, De Wang, Zhiwei Deng, Minghui Fu
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摘要

【摘要】石竹(Eichhornia crassipes)禾苗能有效地从水中吸收氮和磷。克隆crassipes磷酸盐转运蛋白1 (EcPHT1)并研究其特性,有助于解释crassipes吸收和运输无机磷酸盐(Pi)的分子机制,开发新的磷酸盐转运遗传资源,为提高磷酸盐利用效率的作物的遗传育种提供依据。本文利用RACE方法克隆了全长1572 bps的EcPT1,该基因编码523个氨基酸的多肽,包含11个跨膜区,是植物PHT1的典型结构。系统发育分析还表明,EcPT1与其他植物pht1具有密切的亲缘关系。定量实时RT-PCR (qRT-PCR)结果显示,低磷胁迫下,根中EcPT1表达上调。此外,从缺Pi转移到充足Pi后,叶片中EcPT1的表达受到抑制。酵母功能互补实验表明,EcPT1能在175 μM Pi条件下补充磷酸缺陷酵母PHO84的磷酸运输功能,并使其在pH = 5或6时生长最佳。这些结果提示EcPT1可能是一个磷酸转运基因。其在不同组织中的表达不同。
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Cloning and Characterisation of a novel phosphate transporter gene EcPT1 from Eichhornia crassipes
ABSTRACT Eichhornia crassipes (Mart.) Solms can absorb nitrogen and phosphorus efficiently from water. Cloning the phosphate transporter 1 from E. crassipes (EcPHT1) and studying its characteristics can help to explain the molecular mechanism of absorption and transportation of inorganic phosphate (Pi) in E. crassipes and develop new genetic resources for phosphate transport to aid genetic breeding of crops with increased phosphate use efficiency. In this paper, we have used RACE method to clone the EcPT1 with 1572 bps which encoded a polypeptide of 523 amino acids and contained 11 transmembrane regions which was the typical structure of plant PHT1. The phylogenetic analysis also showed EcPT1 was closely related to other plant PHT1s. Quantitative real-time RT-PCR (qRT-PCR) showed that the EcPT1 expression in roots was up-regulated under low phosphorus stress . Moreover the EcPT1 expression in leaves was suppressed after E. crassipes was transferred to sufficient Pi from deficient Pi. Yeast functional complementation tests showed that the EcPT1 could complement the phosphate transport function of the Pi deficient yeast PHO84 in 175 μM Pi and make it grow best in pH = 5 or 6. All these results indicated that EcPT1 might be a phosphate transporter gene. Its expression was different in different tissues.
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