目的 克隆猪苓Polyporus umbellatus菌核无机磷酸盐转运蛋白基因并进行生物信息学和表达模式分析。方法 采用RT-PCR技术从猪苓菌核中克隆得到1个无机磷酸盐转运蛋白（inorganic phosphate transporter）基因，利用生物信息学软件预测蛋白的理化性质、结构域、信号肽和跨膜结构等分子特性；采用Clustal W2以及MEGA 6.0分别进行氨基酸多序列比对和进化关系分析；实时荧光定量PCR分析基因表达模式。结果 猪苓无机磷酸盐转运蛋白基因命名为PuPiT（NCBI登录号：KU179154）。该基因开放读码框全长为1 590 bp，编码530个氨基酸。PuPiT蛋白相对分子质量为57 552，等电点6.82。氨基酸序列分析表明，PuPiT蛋白具有12个跨膜区。氨基酸序列多重比对及系统发育树结果显示PuPiT与Moniliophthora rorer亲缘关系最近，与Moniliophthora roreri、双色蜡蘑Laccaria bicolor、Heterobasidion irregulare有较高的同源性，荧光定量PCR结果表明，PuPiT在与蜜环菌共生的猪苓菌核及未与蜜环菌共生的菌核中都有表达，其中共生部分的表达量显著上调，约是未共生部位的12倍，说明其参与猪苓与蜜环菌共生过程。结论 PuPiT基因克隆和分子特征为进一步研究揭示其在猪苓菌核磷元素转运及与蜜环菌共生过程中的调控作用奠定基础。

Objective To clone an inorganic phosphate transporter (PiT) gene from Polyporus umbellatus and perform the bioinformatics and expression mode analysis. Methods Using RT-PCR.to clone the full-length cDNA of PiT. The characteristics of physiochemical properties, conserved domains, signal peptide and transmembrane domain of the predicted PiT protein were determined by using bioinformatic tools. Results A inorganic phosphate transporter (PiT) gene (NCBI:KU179154), designated as PuPiT, was cloned from Polyporus umbellatus sclerotia by RT-PCR. The full open reading frame cDNA sequence of PuPiT was 1 590 bp, encoding a putative PiT protein with 530 amino acids with a molecular weight of 57 552, and a theoretical pI of 6.82. The amino acids possess 12 membrane-spanning domains. Phylogenetic tree analysis indicated that PuPiT had the highest similarity with PiT from Moniliophthora rorer, and had high similarity with Moniliophthora roreri, Laccaria bicolor, and Heterobasidion irregulare. Quantitative real-time PCR showed that PuPiT expressed in both the symbiotic part and non-symbiotic part. Meanwhile, the expression of PuPiT in the symbiotic part was significantly up-regulated, about 12 times more than that in the non-symbiotic part. This result showed that PuPiT might play an important role in the Pi accumulating. Conclusion Molecular cloning and characterization of the novel PuPiT gene will be useful for further functional determination of the gene involving in phosphorus translocation regulation and symbiotic process.

中国医学科学院医学与健康科技创新工程经费资助（2017-I2M-3-013）；江苏省自然科学基金资助项目（BK20170311）