Cloning, Sequencing and Expression of the Brucella Melitensis Novel Lomr Protein

Abstract

Aim: Brucellosis is a zoonotic infection transmitted from animal to human directly by the animal or indirectly

from their products. It is worldwide distribution especially in the middle east including Iraq and Kurdistan

Region. Materials and methods: The gene coding for the outer membrane protein assembly factor LomR

of 18kDa, now designated of Brucella melitensis 16M was cloned and sequenced. Cloning of insert DNA

from bacteria into pET-28+ allowed the selection of a plasmid bearing a 5.5-kb NcoI fragment that seemed

to contain the entire omp gene under control of its own promoter. Results: LomR was localized within a

region between the NcoI and XhoI insert of approximately 396 bp. The reliability of the constructed plasmid

was established by restriction enzyme mapping and sequencing. LomR was expressed after induction with

IPTG in Escherichia coli BL21. Recombinant LomR was purified by chromatography through Ni-agarose.

Sequencing of this region revealed an open reading frame of 390 bp encoding a protein of 132 amino acids

and a predicted molecular mass of 20 KDa. Conclusion: The availability of recombinant LomR and the

identification of the antigenic determinant recognized will allow the evaluation of their potential protective

activity and their potential for the development of subunit vaccine against brucellosis.