伪狂犬病病毒Ea株gM和gN基因的克隆与结构分析

gM和Ng是最近经经典免疫沉淀法确定的伪狂犬病病毒（PRV）第三对异源糖蛋白二聚体。根据PRV国外Ka株的核苷酸序列，设计两对分别包含gM和Ng完整编码的特异性引物，以PRV国内地方分离株（Ea株）的细胞感染物为模板，PCR扩增出大小约1．2kb和0．3kb的特异性片段。将扩增物克隆到杆状病毒转座载体pEastBacl中，酶切鉴定证实后进行序列测定。结果表明：Ea株gM、Ng基因分别编码394和98个氨基酸，与Ka株的同源性分别为98．4％和97．6％。DNATool和DNASIS软件对gM、Ng进行二级结构预测，发现gM存在8个潜在跨膜区和一个N－糖基化位点，是典型的能反复多次跨膜的Ⅲ型糖蛋白；gN具有N－端信号肽序列、C－端跨膜区和潜在0－糖基化位点，属Ⅰ型糖蛋白。上述结果为下一步深入研究gM、Ng的相互作用位点及二聚体的功能奠定了功能。

Cloning and Identification of Genes Encoding for Glycoprotein M and N of Pseudorabies Virus Strain Ea

Glycoproteins M and N of Pseudorabies virus(PRV)can form a disulfide_linked complex.According to the nucleotide sequences of PV Ka strain,two pairs of primers corresponding to the gM and gN gene were synthesized,and the fragments containing the gM and gN coding domains of PRV Ea strain were amplified by polymerase chain reaction(PCR) from the infectious cell culture.The purified PCR products were inserted into transfer vector pFastBacl under the control of the polyhedrin promoter,resulting in plasmids pFM,pFN and the sequences were obtained by Sanger's sequencing technique.When compared with PRV Ka strain,gM and gN displayede 98.4% and 97.6% animo acid identities.The results of secondary_class analysis showed that gM containing a potencial N_glycosylation sites and eight hydrophobic domains predicted to be transmembrane sequences;gN had a hydrophobic putative signal peptide and transmembrane domains.The above results will lay the foundation for further studying the interacting sites and the funcation of the gM/gN complex.