Molecular cloning,characterization and expression analysis of coagulation factor Ⅱ gene in grass carp(Ctenopharyngodon idella)

【作者】 彭慧珍； 许宝红； 肖调义；

【Author】 Huizhen Peng;Baohon Xu;Tiaoyi Xiao;

【机构】 湖南农业大学动物科学技术学院；

【摘要】 Coagulation factorⅡ,also named prothrombin,is an important factor of exogenous coagulation system.Studies have shown that it is not only involved in blood coagulation process,but also can be induced a series of inflammatory reaction.It has been studied in mammals and other species,but less research of grass carp(Ctenopharyngodon idella),a commercially important freshwater fish found in China.In this study,the full-length c DNA of grass carp coagulation factorⅡ(Gc CFⅡ)was cloned using a RACE-Ready c DNA Kit,grass carp were challenged with Grass carp reovirus873(GCRV873).The temporal expression profiles of Gc CFⅡin the thymus,gills,liver,spleen,and head kidney was detected at0 h,24 h,48 h,72 h,96 h,and 138 h by Real-time q PCR.The results showed that the full-length of Gc CFⅡwas 1718 bp,which consisted of 25 bp 5′UTR,118 bp 3′UTR and 1575 bp ORF,encoding 524 amino acids.It contained four specific hit,Gla,KR,thrombin light chain and Tryp-SPc.The Genbank accession number was KF937388.Phylogenetic analysis suggested that the Gc CFⅡgene evolution was consistent with the evolution of the species.After infected with GCRV873,the Gc CFⅡexpression profiles showed a tendency of fluctuation,the relative expression of Gc CFⅡfalled at first,then increased,and reached to the peak at 72h,and liver and gill were the two most sensitive tissues response to the virus.The results indicated that,Gc CFⅡgene maybe a protein along with the hemorragic disease process,participated in the early acute phase reaction and played an important role in the immune response of grass carp against GCRV873 infection.更多还原

【Abstract】 Coagulation factor Ⅱ,also named prothrombin,is an important factor of exogenous coagulation system.Studies have shown that it is not only involved in blood coagulation process,but also can be induced a series of inflammatory reaction.It has been studied in mammals and other species,but less research of grass carp(Ctenopharyngodon idella),a commercially important freshwater fish found in China.In this study,the full-length c DNA of grass carp coagulation factor Ⅱ(Gc CFⅡ) was cloned using a RACE-Ready c DNA Kit,grass carp were challenged with Grass carp reovirus873(GCRV873).The temporal expression profiles of Gc CFⅡ in the thymus,gills,liver,spleen,and head kidney was detected at 0 h,24 h,48 h,72 h,96 h,and 138 h by Real-time q PCR.The results showed that the full-length of Gc CFⅡ was 1718 bp,which consisted of 25 bp 5′UTR,118 bp 3′UTR and 1575 bp ORF,encoding 524 amino acids.It contained four specific hit,Gla,KR,thrombin light chain and Tryp-SPc.The Genbank accession number was KF937388.Phylogenetic analysis suggested that the Gc CFⅡ gene evolution was consistent with the evolution of the species.After infected with GCRV873,the Gc CFⅡ expression profiles showed a tendency of fluctuation,the relative expression of Gc CFⅡ falled at first,then increased,and reached to the peak at 72 h,and liver and gill were the two most sensitive tissues response to the virus.The results indicated that,Gc CFⅡ gene maybe a protein along with the hemorragic disease process,participated in the early acute phase reaction and played an important role in the immune response of grass carp against GCRV873 infection.更多还原