Li, H.-t., Wang, L., Liu, D. and Yang, X.-q. 2012. Characterization and functional significance of polymorphisms in porcine Toll-like receptor (TLR) 5 gene. Can. J. Anim. Sci. 92: 409-415. Toll-like receptor (TLR) 5 plays an important role in host defenses by recognizing bacterial flagellin and signaling to initiate immune responses. Polymorphisms in the TLR5 gene have a profound influence on receptor function and host susceptibility/resistance to infectious disease, as suggested by studies in humans and other species. In the present study, we characterized polymorphisms and determined their functional significance in the porcine TLR5 gene. Four novel non-synonymous single nucleotide polymorphisms (SNPs), c.176C>T (p.R59M), c.902C>T (p.S301F), c.959T>A (p.F320Y), and c.1796C>T (p.T599M) (reference sequence: GenBank No. AB208697), were first identified by sequencing the complete coding sequences (CDS) of the TLR5 gene in the Min pig, an indigenous Chinese pig breed known for its high general resistance to disease. SNP c.1796C>T (p.T599M) is located in one of the six predicted N-glycosylation sites in the extracellular domain of the TLR5 protein. By measuring protein activation, as represented by nuclear factor κB activity, in transiently transfected PK-15 cells with TLR5 expression vectors carrying site-directed mutations, we demonstrated that the previously discovered SNP c.1205C>T, leading to the amino acid substitution of proline by leucine, attenuated the responses of the receptor to flagellin (P<0.01). Further functional investigation on SNP c.1205C>T is necessary to determine its possible role in disease susceptibility in pigs and may facilitate pig breeding aimed at improving disease resistance.