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东方田鼠对血吸虫具有天然抗性。为筛选和分析东方田鼠抗血吸虫抗性相关基因,以日本血吸虫童虫可溶性裂解物为探针,筛选东方田鼠肝脏噬菌体展示cDNA文库。经三轮筛选,特异性噬菌体得到有效富集(375倍)。随机挑取92个克隆进行序列测定,获得了19条有效EST序列。其中13个条EST序列与已知基因或表达序列标签同源,6个EST序列与已知基因或表达序列标签均无同源性,为新的表达序列标签。将19个EST序列的阳性噬菌体克隆和血吸虫童虫共培养,其中4号(GenBank Accession No.:EW968294)、13号(GenBank Accession No.:EW968303)、14号(GenBank Accession No.:EW968304)、15号(GenBank Accession No.:EW968305)、18号(GenBank Accession No.:EW968308)克隆均诱导了显著的杀虫效果。综合生物信息学分析结果及体外杀伤试验结果,编码CASP8和FADD类似性细胞程序性死亡调节蛋白、α-2-HS-糖蛋白、M4蛋白、具有R3H结构域的一种mRNA结合蛋白以及三种未知蛋白的编码基因(14、15、18号克隆)可能是东方田鼠抗血吸虫病抗性相关基因。为进一步研究东方田鼠抗血吸虫机理奠定了基础。
Eastern voles have natural resistance to schistosomes. To screen and analyze the resistance gene of Microtus fortis, Schistosoma japonicum Schistosoma japonicum soluble lysate was used as a probe to screen the cDNA library of Microtus fortis liver phage. After three rounds of screening, specific phages are efficiently enriched (375-fold). A total of 92 clones were sequenced and 19 valid ESTs were obtained. Among them, 13 ESTs were homologous with known genes or expressed sequence tags, and 6 ESTs were not homologous to known genes or expressed sequence tags, and were new expressed sequence tags. Positive phage clones of 19 EST sequences were co-cultured with Schistosoma japonicum, No. 4 (GenBank Accession No.:EW968294) No. 13 (GenBank Accession No.:EW968303), No. 14 (GenBank Accession No.:EW968304) Clones No. 15 (GenBank Accession No.:EW968305), No. 18 (GenBank Accession No.:EW968308) all induced significant insecticidal effects. Based on the results of bioinformatics analysis and in vitro cytotoxicity test results, we found that CASP8 and FADD-like apoptotic regulatory proteins, α-2-HS-glycoprotein, M4 protein, an mRNA binding protein with R3H domain and three The unidentified protein encoding genes (clone Nos. 14, 15 and 18) may be related to resistance to Schistosoma japonicum. Which laid the foundation for further study on the mechanism of anti-schistosoma japonicum.