中国兽医杂志2024，Vol.60Issue(8)：10-17,8.DOI:10.20157/j.cnki.zgsyzz.2024.08.002
弓形虫srs19d基因敲除株的构建及其在宿主细胞中的生物学功能分析
Construction of Toxoplasma gondii srs19d Gene Knockout Strain and Analysis of Its Biological Functions in Host Cells
摘要
Abstract
To explore the function of a new member of the Toxoplasma gondii surface antigen 1-related sequence(SRS)protein family,SRS19D,this study used the Pru strain of Toxoplasma gondii type Ⅱ as a parent to construct the PruΔsrs19d knockout strain using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9(CRISPR/Cas9)technology.Plaque assays,intracellular proliferation assays,in vitro bradyzoite conversion assays,mouse virulence assays,and mouse brain cyst detection assays were performed.The results from plaque assays and intracellular proliferation assays showed that,compared to the Pru strain,the PruΔsrs19d knockout strain formed significantly smaller plaques on human foreskin fibroblast(HFF)cells(P＜0.05)and produced significantly fewer tachyzoites within the parasitophorous vacuole(P＜0.05).The in vitro bradyzoite conversion assay revealed that the bradyzoite conversion rate of the PruΔsrs19d knockout strain was significantly lower than that of the Pru strain(P＜0.01).The mouse virulence assay showed that the PruΔsrs19d knockout strain did not cause death in ICR mice,in contrast to the Pru strain(P＜0.05).The mouse brain cyst detection assay indicated that the number of brain cysts formed after infection with the PruΔsrs19d knockout strain was significantly reduced compared to the Pru strain(P＜0.01).This study successfully constructed the Toxoplasma gondii PruΔsrs19d knockout strain and discovered that the srs 19d gene plays a crucial role in the parasite's in vitro growth,replication,virulence,and chronic infection,providing a reference for further investigation into the function and mechanism of SRS19D.关键词
弓形虫/表面抗原1相关序列(SRS)/srs19d/成簇规律间隔短回文重复-CRISPR相关蛋白9(CRISPR/Cas9)/生物学功能Key words
Toxoplasma gondii/surface antigen 1-related sequences(SRS)/srs19d/CRISPR/Cas9/biological function分类
农业科技引用本文复制引用
牛水珠,潘明,葛层层,黄思扬..弓形虫srs19d基因敲除株的构建及其在宿主细胞中的生物学功能分析[J].中国兽医杂志,2024,60(8):10-17,8.基金项目
江苏省杰出青年基金项目(BK20190046) （BK20190046）
江苏高校优势学科建设工程资助项目(PAPD) （PAPD）
