• Vaccine Name: B. abortus DNA vaccine encoding RplL and Omp16
• Target Pathogen: Brucella spp.
• Target Disease: Brucellosis
• Vaccine Ontology ID: VO_0011380
• Type: DNA vaccine
• Status: Research
• Antigen: B. abortus ribosomal protein rplL and outer membrane lipoprotein 16
• RplL gene engineering:
  • Type: DNA vaccine construction
  • Description: Full-length open reading frames of the L7/L12 gene and Omp16 gene were amplified with PCR from the genome of attenuated B. abortus strain RB51. The gene amplified with L7/L12 primers (FL and RL-1) and the gene amplified with Omp16 primers (FO and RO) were inserted into pcDNA3.1(+) vector (Invitrogen) at the EcoRV/XhoI and BamHI/XhoI sites to construct recombinant plasmids L7/L12-pcDNA3.1 and Omp16-pcDNA3.1, respectively. To construct the recombinant fusion plasmid L7/L12-Omp16-pcDNA3.1, the L7/L16 gene fragment was amplified with the L7/L16 PCR primers (FL and RL-2) first, which removed only the TAA stop codon from the L7/L16 gene. This PCR product and the amplified Omp16 gene above were digested with EcoRV/BamHI and BamHI/XhoI, respectively, and ligated with T4 ligase; the ligated product was then inserted into the pcDNA3.1(+) vector between the EcoRV and XhoI sites (Luo et al., 2006b).
  • Detailed Gene Information: Click Here.
• Omp16 gene engineering:
  • Type: DNA vaccine construction
  • Description: Full-length open reading frames of the L7/L12 gene and Omp16 gene were amplified with PCR from the genome of attenuated B. abortus strain RB51. The PCR primers were designed as shown in Table 1. The gene amplified with L7/L12 primers (FL and RL-1) and the gene amplified with Omp16 primers (FO and RO) were inserted into pcDNA3.1(+) vector (Invitrogen) at the EcoRV/XhoI and BamHI/XhoI sites to construct recombinant plasmids L7/L12-pcDNA3.1 and Omp16-pcDNA3.1, respectively. To construct the recombinant fusion plasmid L7/L12-Omp16-pcDNA3.1, the L7/L16 gene fragment was amplified with the L7/L16 PCR primers (FL and RL-2) first, which removed only the TAA stop codon from the L7/L16 gene. This PCR product and the amplified Omp16 gene above were digested with EcoRV/BamHI and BamHI/XhoI, respectively, and ligated with T4 ligase; the ligated product was then inserted into the pcDNA3.1(+) vector between the EcoRV and XhoI sites (Luo et al., 2006b).
  • Detailed Gene Information: Click Here.
• DNA vaccine plasmid:
  • DNA vaccine plasmid name:
  • DNA vaccine plasmid VO ID: VO_0000158
• Immunization Route: Intramuscular injection (i.m.)

Mouse Response

• Host Strain: BALB/c
• Vaccination Protocol: The mice were anesthetized with methoxyflurane (Metofan; Mallinckrodt) and inoculated intramuscularly with 100 μg of pcDNA3.1-L7/L12-Omp16, pcDNA3.1-L7/L12, or pcDNA3.1-Omp16 in 100 μl of PBS (50 μl of the solution was injected into each tibialis anterior muscle). The control mice were infected with PBS or the expression vector alone (pcDNA3.1). Each mouse in another group was injected with 10 μg of rL7/L12-Omp16 in 100 μl PBS according to the same schedule. Each mouse was injected on weeks 0, 2, and 4. The mice used as positive controls were inoculated intraperitoneally on day 0 with 2× 10^8 CFU of B. abortus strain RB51 in 0.2 ml of PBS (Luo et al., 2006b).
• Challenge Protocol: Two weeks after the final vaccination, five mice from each group were challenged intraperitoneally with relatively higher dose of strain 544 (5 × 10^5 CFU). Four weeks postchallenge, the mice were killed by cervical dislocation, and their spleens were removed aseptically and weighed (Luo et al., 2006b).
• Efficacy: This divalent DNA vaccine induced a significant level of protection against challenge with the virulent B. abortus in BALB/c mice (Luo et al., 2006b).
• Host Ighg1 response
  • Description: Sera collected 2 weeks after the last immunization were assayed for the presence of L7/L12- and/or Omp16-specific antibodies by ELISA. The pcDNA 3.1-L7/L12-OMP16 vaccine elicited significantly higher levels of IgG1 antibodies than did the negative control vaccine, pcDNA 3.1 (Luo et al., 2006b).
  • Detailed Gene Information: Click Here.
• Host Ighv1-9 response
  • Description: Sera collected 2 weeks after the last immunization were assayed for the presence of L7/L12- and/or Omp16-specific antibodies by ELISA. The pcDNA 3.1-L7/L12-OMP16 vaccine elicited significantly higher levels of IgG2a antibodies than did the negative control vaccine, pcDNA 3.1 (Luo et al., 2006b).
  • Detailed Gene Information: Click Here.