短柄草磷转运蛋白家族基因的克隆及表达分析

Cloning and Expression Analysis of Phosphate Transporters Pht1 in Brachypodium distachyon

植物高亲和力磷转运蛋白Pht1家族是一类H2PO4-/H＋共转运子,主要在根系中负责磷的吸收和转运,其表达受低磷调控,对该家族成员的研究有助于揭示磷的吸收和转运机制。根据水稻、拟南芥、大麦中磷转运蛋白基因的序列,预测出短柄草Pht1家族共有12个成员,分别命名为BRAdi;Pht1;1~BRAdi;Pht1;12,设计基因特异引物,对基因组和cDNA全长基因进行克隆、测序,通过对基因编码的氨基酸序列同源比对分析,构建了系统发生树,并利用实时荧光定量PCR技术对各基因成员的表达模式进行了分析。结果表明,预测到的成员具有Pht1家族的典型结构,成员间的同源性高,进化树分析将这12个同源基因分为不同的亚组,这些同源基因与大麦的同源性较高,其次是水稻,而与拟南芥的同源性最低。这些基因在不同的组织中表达量不同,在种子中的表达量最高,有5个基因在苗期根中的表达显著高于叶片。

High-affinity phosphate transporter family（Pht1） belongs to a type of H2PO4-/H＋ symporters,mainly responds to the acquisition and translocation of phosphorus in root,and its expression is regulated by phosphorus deficiency.Investigation of this gene family will be helpful to unravel the mechanism of phosphate uptake and translocation in plant.According to the gene sequences of Pht1 in rice,Arabidopsis and barley,and the whole genome sequences of Brachypodium distachyon,12 Pht1 members were predicted by Blastn analysis,which were subsequently denominated as BRAdi;Pht1;1~BRAdi;Pht1;12.The gene and genomic sequences of each Pht1 members were obtained by PCR and cloning with gene specific primers using genomic DNA and cDNA as the template.The amino acid sequences of all the members were aligned,its phylogenetic relationship was analyzed,and gene expression pattern was investigated with quantitative realtime PCR.The results showed that all the predicted members have typical conserved structures of Pht1 gene and they are highly homological to each other.The cloned Pht1 members were divided into several subgroups by phylogenetic analysis,which revealed that Pht1 members from Brachypodium distachyon are highly homological to those from barley,but less homological to those from rice and Arabidopsis.The expression patterns also revealed that these Pht1 genes were differentially expressed in tissues,of which five genes have higher expression level in root samples than that in leaves of young seedlings.