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Production of recombinant capsid protein of Macrobrachium rosenbergii nodavirus (r‐ MCP 43) of giant freshwater prawn, M. rosenbergii (de Man) for immunological diagnostic methods
Author(s) -
Farook M A,
Madan N,
Taju G,
Abdul Majeed S,
Nambi K S N,
Sundar Raj N,
Vimal S,
Sahul Hameed A S
Publication year - 2014
Publication title -
journal of fish diseases
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.819
H-Index - 85
eISSN - 1365-2761
pISSN - 0140-7775
DOI - 10.1111/jfd.12156
Subject(s) - macrobrachium rosenbergii , prawn , antiserum , biology , western blot , capsid , recombinant dna , virology , microbiology and biotechnology , dot blot , virus , antibody , gene , fishery , immunology , biochemistry
Abstract White tail disease (WTD) caused by Macrobrachium rosenbergii nodavirus ( Mr NV) and extra small virus (XSV) is a serious problem in prawn hatcheries. The gene for capsid protein of Mr NV (MCP43) was cloned into pRSET B expression vector. The MCP43 protein was expressed as a protein with a 6‐histidine tag in Escherichia coli GJ1158 with NaCl induction. This recombinant protein, which was used to raise the antiserum in rabbits, recognized capsid protein in different WTD‐infected post‐larvae and adult prawn. Various immunological methods such as Western blot, dot blot and ELISA techniques were employed to detect Mr NV in infected samples using the antiserum raised against recombinant MCP43 of Mr NV. The dot blot assay using anti‐ rMCP 43 was found to be capable of detecting Mr NV in WTD‐infected post‐larvae as early as at 24 h post‐infection. The antiserum raised against r‐MCP43 could detect the Mr NV in the infected samples at the level of 100 pg of total protein. The capsid protein of Mr NV estimated by ELISA using anti‐ rMCP 43 and pure r‐MCP43 as a standard was found to increase gradually during the course of infection from 24 h p.i. to moribund stage. The results of immunological diagnostic methods employed in this study were compared with that of RT‐PCR to test the efficiency of antiserum raised against r‐MCP43 for the detection of Mr NV. The Western blot, dot blot and ELISA detected all Mr NV‐positive coded samples as detected by RT‐PCR.