CAJ | 학술논문

利用鲤(Cyprinus carpio)上皮细胞(epitheliaoma papulosum cyprini, EPC)培养传染性造血器官坏死病毒-Sn1203分离株(IHNV-Sn1203),根据GenBank中IHNV G蛋白基因开放阅读框(open reading frame, ORF)的序列设计引物(GenBank序列编号AB288207),采用RT-PCR的方法克隆得到IHNV-Sn1203株G蛋白全长ORF,克隆至表达载体pET27b(+)中,构建了pET27-G重组质粒,并进行了测序分析。生物信息学分析结果显示, IHNV-Sn1203株G蛋白基因序列长度为1527 bp,与韩国株具有最高的核酸同源性(96.86%)和氨基酸同源性(97.05%)。该基因编码508个氨基酸残基,推导分子量约为56.55 kD,等电点为6.15；氨基酸序列分析表明, G蛋白富含丝氨酸、苏氨酸和酪氨酸,存在28个潜在的磷酸化位点；存在4个潜在的N-糖基化位点和7个潜在的O-糖基化位点；G蛋白N端含有20个氨基酸的信号肽；亲水性大于输水性；位于483~508位氨基酸存在一跨膜区；抗原表位预测显示抗原性良好；系统进化树分析显示, IHNV-Sn1203株与日本株和韩国株聚为一簇,都属于JRt基因型。
이용리(Cyprinus carpio)상피세포(epitheliaoma papulosum cyprini, EPC)배양전염성조혈기관배사병독-Sn1203분리주(IHNV-Sn1203),근거GenBank중IHNV G단백기인개방열독광(open reading frame, ORF)적서렬설계인물(GenBank서렬편호AB288207),채용RT-PCR적방법극륭득도IHNV-Sn1203주G단백전장ORF,극륭지표체재체pET27b(+)중,구건료pET27-G중조질립,병진행료측서분석。생물신식학분석결과현시, IHNV-Sn1203주G단백기인서렬장도위1527 bp,여한국주구유최고적핵산동원성(96.86%)화안기산동원성(97.05%)。해기인편마508개안기산잔기,추도분자량약위56.55 kD,등전점위6.15；안기산서렬분석표명, G단백부함사안산、소안산화락안산,존재28개잠재적린산화위점；존재4개잠재적N-당기화위점화7개잠재적O-당기화위점；G단백N단함유20개안기산적신호태；친수성대우수수성；위우483~508위안기산존재일과막구；항원표위예측현시항원성량호；계통진화수분석현시, IHNV-Sn1203주여일본주화한국주취위일족,도속우JRt기인형。
The glycoprotein (G) is one of the structural proteins on the surface of infectious haematopoietic necrosis virus (IHNV) and has important functions. In this study, the open reading frame of the glycoprotein gene was amplified by reverse transcriptase-PCR (RT-PCR) from isolate IHNV-Sn1203 and cloned into the pET27b(+) vector. The structure and characteristics of the glycoprotein gene were analyzed using bioinformatics software. The results showed that the G gene was 1 527 bp, encoding a 508 amino acid protein, Isolate IHNV-Sn1203 shared closer identity with strains isolated from South Korea, and the homology of at nucleotide and amino acid level of the G gene were 96.68%and 97.05%, respectively. The molecular weight was 56.55 kD and the isoelectric point was 6.15. The glycoprotein was rich inserine, threonine and tyrosine, and contained 28 potential phosphorylation sites. According to the protein structure prediction, the glycoprotein may have four N-glycosylationsites and seven O-glycosylation sites. The glycoprotein is hydrophilic, with an N-terminal signal peptide and a transmembrane region from amino acid 483 to 508. The glycoprotein was esti-mated to be highly antigenic. Phylogenetic analysis showed that isolate IHNV-Sn1203 was in the same branch as iso-lates from Japan and South Korea, belonging to the JRt genotype. This study established a foundation for research into the genetic background, pathogenesis, and molecular epidemiology of IHNV.