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Induction, purification and partial characterization of vitellogenin in an I ndian major carp C atla catla ( H am.)
Author(s) -
Jena Bipulendu,
Mohanty Jyotirmaya,
Das Radha C,
Garnayak Sushil K,
Nandi Samiran
Publication year - 2013
Publication title -
aquaculture research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.646
H-Index - 89
eISSN - 1365-2109
pISSN - 1355-557X
DOI - 10.1111/j.1365-2109.2012.03195.x
Subject(s) - vitellogenin , catla , biology , polyclonal antibodies , carp , size exclusion chromatography , chromatography , western blot , microbiology and biotechnology , antibody , biochemistry , fish <actinopterygii> , chemistry , fishery , immunology , enzyme , gene
Vitellogenin was purified from the serum of 17‐β estradiol (E 2 )‐induced juvenile C atla catla using a simple two‐step purification procedure i.e. selective chemical precipitation followed by gel filtration chromatography. Purified protein migrated as single band in a native gradient PAGE which indicated the purity of the sample. The molecular weight of the native catla vitellogenin (~440 kDa) was determined using gel filtration chromatography. In SDS ‐ PAGE under reducing conditions catla vitellogenin dissociated into three major sub units at 115 kDa, 102 kDa and 73 kDa along with a few faint bands. Confirmation of purified protein as catla vitellogenin was supported by multiple physiological evidences, e.g. absence in male as well as juvenile sera and presence in matured female fish, ability to be synthesized upon estradiol injection in immature fish and certain unique biochemical properties like high molecular weight, phospholipoglycoprotein nature of the molecule. Western blot analysis showed that polyclonal antibody raised against purified protein detected vitellogenin in the sera of catla and in a few species selected from C yprinidae family. These antisera were used to detect vitellogenin in liver tissue of hormone‐induced catla using immunohistochemistry and its applicability in other immunoassays is discussed.