title: Prokaryotic Expression ofα-13 Giardin Gene and Its Intracellular Localization inGiardia lamblia.
abstract: To study prokaryotic expression and subcellular localization ofα-13 giardin inGiardia lambliatrophozoites,α-13 giardin gene was amplified and cloned into prokaryotic expression vector pET-28a(+). The positive recombinant plasmid was transformed intoE. coliBL21(DE3) for expression by using IPTG and autoinduction expression system (ZYM-5052). The target protein was validated by SDS-PAGE and Western blotting and purified by Ni-NTA Resin. Rabbits were immunized with purified fusion proteins for preparation of polyclonal antibody; then the intracellular location ofα-13 giardin was determined by fluorescence immunoassay. The results showed that the length ofα-13 giardin gene was 1038 bp, encoding a polypeptide of 345 amino acids. The expressed product was a fusion protein with about 40 kDa largely present in soluble form. The target protein accounted for 21.0% of total proteins after being induced with IPTG, while it accounted for 28.8% with ZYM-5052. The anti-α13-giardin polyclonal antibody possessed good antigenic specificity as well as excellent binding activity with recombinantα-13 giardin. Immunofluorescence assays revealed thatα-13 giardin was localized in the cytoplasm ofG. lambliatrophozoite, suggesting that it is a cytoplasm-associated protein. The present study may lay a foundation for further functional research onα-13 giardin ofG. lamblia.
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Xingang
authors.initials: X
authors.lastname: Yu
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Auwalu Yusuf
authors.identifier: 0000-0003-2627-1389
authors.initials: AY
authors.lastname: Abdullahi
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Sheng
authors.identifier: 0000-0003-1663-1379
authors.initials: S
authors.lastname: Wu
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Weida
authors.initials: W
authors.lastname: Pan
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Xianli
authors.initials: X
authors.lastname: Shi
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hu
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Liping
authors.initials: L
authors.lastname: Tan
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Kangxin
authors.initials: K
authors.lastname: Li
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510542, China.
authors.forename: Guoqing
authors.identifier: 0000-0001-9393-1573
authors.initials: G
authors.lastname: Li
mesh_terms: D000818:Animals; D003593:Cytoplasm / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D016829:Giardia lamblia / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015800:Protozoan Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D053058:Trophozoites / Q000737:chemistry / Q000378:metabolism
chemical_list: D003598:Cytoskeletal Proteins; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C038207:giardin protein, Giardia lamblia