Construction of Larval cDNA Expression Library and Immunological Identification of Positive Clones in Tick Haemaphysalis qinghaiensis  

作　　者：ZHAO Hai-ping YIN Hong LI Chun-yi GAO Jin-liang LI You-quan GUAN Gui-quan LIU Zhi-jie LIU Jun-long YANG Fu-he XING Xiu-mei LUO Jian-xun 

机构地区：[1]State Key Laboratory on Veterinary Etiology （Key Laboratory of Veterinary Parasitology in Gansu Province）, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, P.R.China [2]Institute of Special Wild Economic Animal and Plant Science, Chinese Academy of Agricultural Sciences, Jilin 132109, P.R.China [3]AgResearch Invermay Agricultural Centre, Mosgiel Private Bag 50034, New Zealand

出　　处：《Agricultural Sciences in China》2010年第6期896-902,共7页中国农业科学（英文版）

基　　金：supported by the Na- tional High-Tech R&D Program (2006AA10A207);the National Key Technology R&D Program (2007- BAD40B06);the Natural Resource Platform Project (2005DKA21104);the National Natural Science Foundation of China (30270992) as well

摘　　要：A primary cDNA expression library with a titer of 5.0 × 105 PFU mL-1 was constructed from mRNA extracted from larval Haemaphysalis qinghaiensis ticks in order to identify certain genes,which would then be used as candidate molecules for development of effective vaccines to control this parasite.Totally 11 positive clones,which designated as HqL01-11,were obtained by immunoscreening of the library using a polyclonal antibody generated in rabbit with larval tick protein extract.Results of sequence analysis from BLASTN searching revealed that 6 of them had no significant homology with the adult H.qinghaiensis ticks’ known genes,4 of them had no significant homology with all genes deposited in GenBank database.HqL07,HqL08,HqL09,and HqL11 were deposited to GenBank database,and accession numbers were EF605263,EF605264,EF605265,and EF605266,respectively.Subsequently,HqL07 and HqL09 were expressed in vitro and the molecular weights of the corresponding expressed products were 60 and 70 kDa,respectively.Western blot analyses showed that HqL07 and HqL09 had immunogenicity.This study laid the foundation for future production of genetically engineered vaccines for the immunological control of H.qinghaiensis.A primary cDNA expression library with a titer of 5.0 × 105 PFU mL-1 was constructed from mRNA extracted from larval Haemaphysalis qinghaiensis ticks in order to identify certain genes,which would then be used as candidate molecules for development of effective vaccines to control this parasite.Totally 11 positive clones,which designated as HqL01-11,were obtained by immunoscreening of the library using a polyclonal antibody generated in rabbit with larval tick protein extract.Results of sequence analysis from BLASTN searching revealed that 6 of them had no significant homology with the adult H.qinghaiensis ticks’ known genes,4 of them had no significant homology with all genes deposited in GenBank database.HqL07,HqL08,HqL09,and HqL11 were deposited to GenBank database,and accession numbers were EF605263,EF605264,EF605265,and EF605266,respectively.Subsequently,HqL07 and HqL09 were expressed in vitro and the molecular weights of the corresponding expressed products were 60 and 70 kDa,respectively.Western blot analyses showed that HqL07 and HqL09 had immunogenicity.This study laid the foundation for future production of genetically engineered vaccines for the immunological control of H.qinghaiensis.

关 键 词：larval tick Haemaphysalis qinghaiensis cDNA expression library IMMUNOSCREENING immunologicalidentification 

分 类 号：Q969.91[生物学—昆虫学] S852.734[农业科学—基础兽医学]