Expression and purification of a gene encoding a 9.7 kDa PE protein of Mycobacterium avium subsp. paratuberculosis.
Mycobacterium avium subsp. paratuberculosis (Map) contains PE family antigens which are Proline and glutamic acid rich and may play important role as T-cell antigens. In the present study, the Map 1507 ORF encoding 9.7 kDa PE protein was amplified by polymerase chain reaction and cloned into E. coli vector pQE30 UA. The recombinant plasmid designated as pQ PE was transformed into E. coli M15 cells and induced with IPTG revealed the high level expression of 11.9 kDa His-fusion protein as estimated by migration in 15 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Recombinant PE protein was purified by Ni-NTA agarose chromatography. Polyclonal antibodies raised against purified recombinant PE protein reacted with expressed PE protein as well as with Map sonicate. The recombinant PE protein was also recognized by serum from goat with clinical paratuberculosis. The protein elicited significant delayed type hypersensitivity (DTH) skin reaction in mice sensitized with Map. The results indicated that the recombinant PE protein of Map was associated with T-cell response.
Division of Veterinary Biotechnology, ICAR-Indian Veterinary Research Institute, Izatnagar, 243122, Uttar Pradesh, India.
S Chandra
SC
Sekar
Division of Veterinary Biotechnology, ICAR-Indian Veterinary Research Institute, Izatnagar, 243122, Uttar Pradesh, India. ppgnbc@yahho.co.in.
P P
PP
Goswami
Division of Veterinary Biotechnology, ICAR-Indian Veterinary Research Institute, Izatnagar, 243122, Uttar Pradesh, India.
R
R
Deb
DTH; Map; PE; Purification; pQE30 UA