title: Prokaryotic expression of the chicken xanthine oxidase (XOD) subunit and its localization in liver and kidney.
abstract: Xanthine oxidase (XOD) is the members of the molybdenum hydroxylase flavoprotein family and it plays a vital role in the body's purine catabolism. In this study, we cloned the XOD 37kDa subunit protein by using RT-PCR and pMD-18-T clone vector based on the total RNA extracted from chicken liver. The cloning XOD subunit protein gene was ligated into the pET-32a to construct the recombinant plasmid pET-XOD. After the pET-XOD expression vector was transformed into host cells Rosetta (DE3), the recombinant XOD subunit proteins (54.8kDa) were successfully induced by isopropy1 β-d-thiogalactoside (IPTG). Rabbit antiserums were produced by using the purification of the recombinant XOD subunit protein as antigen. The titer of the antiserum was more than 1:102,400 determined by using ELISA. The result of Western blot demonstrated that the antiserum could specifically recognize the chicken liver XOD. Immunohistochemistry and immunofluorescence showed that the XOD mainly presented in the cytoplasm of chicken hepatocytes and proximal tubular epithelial cells. Our results indicated that the XOD subunit protein polyclonal antibody prepared by this method could be used for the further researches of the biological function of the XOD in the chicken.
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Huayuan
authors.initials: H
authors.lastname: Lin
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Yanjun
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Qiqi
authors.initials: Q
authors.lastname: Huang
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China. Electronic address: Xqguo20720@aliyun.com.
authors.forename: Xiaoquan
authors.initials: X
authors.lastname: Guo
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China. Electronic address: pingliujx@163.com.
authors.forename: Ping
authors.initials: P
authors.lastname: Liu
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Weilian
authors.initials: W
authors.lastname: Liu
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Caiying
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Huabin
authors.initials: H
authors.lastname: Cao
authors.affiliation: Clinical Veterinary Laboratory, College of Animal Science and Technology, Jiangxi Agricultural University, Jiangxi, China.
authors.forename: Guoliang
authors.initials: G
authors.lastname: Hu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002645:Chickens / Q000235:genetics; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005818:Genetic Engineering; D005822:Genetic Vectors / Q000235:genetics; D007668:Kidney / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D021382:Protein Sorting Signals; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D014969:Xanthine Oxidase / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*
chemical_list: D000939:Epitopes; D021382:Protein Sorting Signals; D021122:Protein Subunits; D014969:Xanthine Oxidase
keywords: Chicken; ELISA; Immunofluorescence; Immunohistochemistry; Western blot; Xanthine oxidase