CAJ | 학술논문

根据已发表的柔嫩艾美耳球虫（Houghton 株）基因序列，设计一对特异性引物，通过 RT-PCR扩增柔嫩艾美耳球虫长春株 SAG2基因，将扩增的片段连入 pMD18T 载体，筛选阳性克隆进行测序和序列分析。结果表明，成功克隆了柔嫩艾美耳球虫长春株 SAG2基因，测得 ORF 全长813 bp；与 GenBank 中收录的 Houghton 株柔嫩艾美耳球虫的 SAG2基因序列相比，有4个碱基发生变异，两者的核苷酸序列同源性为99．50％；柔嫩艾美耳球虫长春株 SAG2基因的推导氨基酸序列同源性为99．63％，其中，A185G 变异导致了编码氨基酸由谷氨酸变为甘氨酸（E62G），A247T 变异导致了编码氨基酸由蛋氨酸变为亮氨酸（M83L），而其他核苷酸的突变为无义突变；SAG2抗原的 N 端和 C 端疏水性较强，并且 N 端前23个氨基酸为跨膜信号肽区，而中间有许多亲水性区域，且显示很强的抗原性。柔嫩艾美耳球虫长春株与 Houghton 株的 SAG2基因编码蛋白相似性很高，具有很强的抗原性，可以作为球虫疫苗的候选抗原来进一步研究。
근거이발표적유눈애미이구충（Houghton 주）기인서렬，설계일대특이성인물，통과 RT-PCR확증유눈애미이구충장춘주 SAG2기인，장확증적편단련입 pMD18T 재체，사선양성극륭진행측서화서렬분석。결과표명，성공극륭료유눈애미이구충장춘주 SAG2기인，측득 ORF 전장813 bp；여 GenBank 중수록적 Houghton 주유눈애미이구충적 SAG2기인서렬상비，유4개감기발생변이，량자적핵감산서렬동원성위99．50％；유눈애미이구충장춘주 SAG2기인적추도안기산서렬동원성위99．63％，기중，A185G 변이도치료편마안기산유곡안산변위감안산（E62G），A247T 변이도치료편마안기산유단안산변위량안산（M83L），이기타핵감산적돌변위무의돌변；SAG2항원적 N 단화 C 단소수성교강，병차 N 단전23개안기산위과막신호태구，이중간유허다친수성구역，차현시흔강적항원성。유눈애미이구충장춘주여 Houghton 주적 SAG2기인편마단백상사성흔고，구유흔강적항원성，가이작위구충역묘적후선항원래진일보연구。
According to the SAG2 gene sequence of E.tenella published in GenBank,a pair of specific prim-ers was designed.Then,the SAG2 gene in E.tenella Changchun strain was amplified by RT-PCR and li-gated into pMD18T vector,the positive clone was sequenced.The results showed that the ORF of E. tenella of Changchun strain SAG2 gene is 813 bp,compared with the published SAG2 gene (Houghton strain)sequence,four nucleotide mutation sites appeared in the SAG2 gene of E.tenella Changchun strain.There is 99.50% homology in nucleotide sequence,and 99.63% homology in its encoding region sequence.Among these mutation sites,A185G mutation led to substitution of glutamic acid by glycine, and A247T mutation led to substitution of methionine by leucine.There is strong hydrophobic at N-termi-nal and C-terminal of SAG2,and there is 23 amino acid transmembrane signal peptide at N-terminal,how-ever,there are a lot of hydrophilic zones in the middle portion,and showed strong antigenicity.These suggest that there is high homology between Changchun strain and Houghton strain of E.tenella SAG2 gene,the protein encoded by SAG2 gene in E.tenella Changchun strain had strong antigenicity,it can be used as a candidate antigen for vaccine development.