Unfortunately, these methods entail eventual inconveniences that limit the practical usefulness of the tagging deletion such as: (a) an excessive attenuation and/or loss of vaccine efficacy (Barrio et?al., 2009; Gonzlez et?al., 2008); TEF2 (b) the lack of specificity of the associated assessments to detect specific antibodies against the deleted antigen in infected animals Cucurbitacin B (Grill et?al., 2009); and (c) a potential positive selective advantage for virulent bacteria against the attenuated vaccine strain (Moreno, 2014). An alternative approach has been the inclusion of antigens xenogenic for immunogenic proteins (Comerci, Pollevick, Vigliocco, Frasch, & Ugalde, 1998; Pollevick, Affranchino, Frasch, & Sanchez, 1991) or the Green Fluorescent Protein (GFP) from your jellyfish (Chacn\Daz et?al., 2011) in combination with associated diagnostic tests. with recombinant GFP induced an intense and long\lasting ( 9?months) Cucurbitacin B anti\GFP serological response readily detectable by the ELISA\GFP. Overall, our results confirm that Rev1 GFP\tagging can be a suitable alternative for identifying vaccinated sheep in infected contexts. species that affects a large variety of domestic and wildlife hosts, including humans (Moreno & Moriyn, 2006). is the main agent of human brucellosis, which is usually related directly to contamination in sheep and goats. The disease induces abortions and reproductive disorders in small ruminants, causing important economic and public health problems (Corbel, 2006; Dean, Crump, Greter, Schelling, & Zinsstag, 2012; Zinsstag, Schelling, Solera, Blasco, & Moriyn, 2011). Due to the absence of suitable vaccines for humans, prevention depends on the control/eradication of the disease in animals. Vaccination, combined or not with test & slaughter, is the only practical strategy to control this important zoonosis in most affected areas (Blasco, 1997; Blasco & Molina\Flores, 2011). Rev1 is the only vaccine available against sheep and goat brucellosis (OIE, 2016). The protective efficacy of this live vaccine is based on its residual virulence, a critical property required to induce a long\lasting Cucurbitacin B effective immunity against Cucurbitacin B field infections (Bosseray & Plommet, 1990). However, Rev1 vaccination induces anti\easy lipopolysaccharide (S\LPS) antibodies that are indistinguishable of those generated after field infections, leading to interferences in standard anti\LPS assessments (Ducrotoy, Conde\lvarez, Blasco, & Moriyn, 2016). Vaccination of exclusively young replacements minimizes this interference (particularly when Cucurbitacin B Rev1 is applied by the conjunctival route) but does not fully solve the problem of differentiating vaccinated animals (Fensterbank, Pardon, & Marly, 1982, 1985). Several approaches to generate vaccines or strategies to allow differentiation between infected and vaccinated animals (DIVA) have been analyzed (Blasco, Moreno, & Moriyn, 2016; Yang et?al., 2013). Since most infected animals react against cell envelope antigens, efforts have been focused on the removal of immunogenic relevant proteins such as outer membrane and binding proteins (Cloeckaert et?al., 2004; Grill et?al., 2009; Guilloteau et?al., 2006; Jacques et?al., 2007) or O\polysaccharide epitopes (Godfroid et?al., 2000; Gonzlez et?al., 2008). Regrettably, these methods entail eventual inconveniences that limit the practical usefulness of the tagging deletion such as: (a) an excessive attenuation and/or loss of vaccine efficacy (Barrio et?al., 2009; Gonzlez et?al., 2008); (b) the lack of specificity of the associated assessments to detect specific antibodies against the deleted antigen in infected animals (Grill et?al., 2009); and (c) a potential positive selective advantage for virulent bacteria against the attenuated vaccine strain (Moreno, 2014). An alternative approach has been the inclusion of antigens xenogenic for immunogenic proteins (Comerci, Pollevick, Vigliocco, Frasch, & Ugalde, 1998; Pollevick, Affranchino, Frasch, & Sanchez, 1991) or the Green Fluorescent Protein (GFP) from your jellyfish (Chacn\Daz et?al., 2011) in combination with associated diagnostic assessments. Both approaches were performed by encoding the xenogenic antigens through a non\integrative plasmid, limiting the usefulness of these vaccines in field conditions. We have developed a Rev1 vaccine strain transporting the gene stably inserted in the chromosome (Rev1::Rev1 vaccine reference strainIdAB collectionRev1::inserted into the site in the intergenic chromosomal regionKm50 r This workRev1pGFPRev1 transporting the non\integrative plasmid pBBR1\2\expressionIdAB collectionH38::Gm biovar 1 transporting mini\TnS17 (pir) transporting the pUC18T\mini\TnP1::P1\fragment.(Choi & Schweizer, 2006)TOP10F/pCR2.1\TOP10F carrying the pCR2.1\TOPO TA Cloning kit (Thermo Fisher Scientific) containing P1\P1\from pCR2.1\S17 (pir) carrying the pUC18R6KT\mini\TnP1::P1\in the intergenic region between and of the chromosome.This workSM10 (pir)/pTNS2 SM10 (pir) carrying the plasmid.