A novel genetically engineered subunit vaccine for Mycoplasma gallisepticum
A technology of mycoplasma gallisepticum and amino acids, which is applied in the direction of vaccines, veterinary vaccines, antibody medical components, etc., can solve the problems of lack, weak immune protection of vaccines, and easy mutation of antigenic proteins, so as to achieve sufficient supply and protein immunogen Good sex and good immunogenicity
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Embodiment 1
[0143] Example 1 Construction and identification of transfer vector pF-MGC1
[0144] 1. MGC1 gene amplification and purification The codon-optimized MGC1 gene (SEQ ID NO: 1) was synthesized in Nanjing GenScript Company and cloned into the pUC17 vector to obtain the pUC-MGC1 plasmid vector. The pUC-MGC1 plasmid was used as a template, and MGC1-F and MGC1-R were used as upstream and downstream primers for PCR amplification (the gene sequences of MGC1-F and MGC1-R are shown in SEQ ID NO. 11 and 12). The amplification system is shown in Table 1.
[0145] Table 1 MGC1 gene amplification system
[0146]
[0147] The reaction conditions were: pre-denaturation at 95°C for 5 minutes; denaturation at 94°C for 45 seconds, annealing at 54°C for 45 seconds, extension at 72°C for 1 minute, 35 cycles; extension at 72°C for 10 minutes.
[0148] The PCR product is subjected to gel electrophoresis to verify the size of the target gene, such as figure 1 As shown, the target band appeared a...
Embodiment 2
[0161] Example 2 Construction and identification of transfer vector pF-MGC2
[0162] 1. MGC2 gene amplification and purification The codon-optimized MGC2 gene (SEQ ID NO: 3) was synthesized in Nanjing GenScript Company and cloned into the pUC17 vector to obtain the pUC-MGC2 plasmid vector. The pUC-MGC2 plasmid was used as a template, and MGC2-F and MGC2-R were used as upstream and downstream primers for PCR amplification (the gene sequences of MGC2-F and MGC2-R are shown in SEQ ID NO. 13 and 14). The amplification system is shown in table 5.
[0163] Table 5 MGC2 gene amplification system
[0164]
[0165]
[0166] The reaction conditions were: pre-denaturation at 95°C for 5 minutes; denaturation at 94°C for 45 seconds, annealing at 54°C for 45 seconds, extension at 72°C for 1 minute, 35 cycles; extension at 72°C for 10 minutes.
[0167] The PCR product is subjected to gel electrophoresis to verify the size of the target gene, such as Figure 4 As shown, the target ba...
Embodiment 3
[0179] Example 3 Construction and identification of transfer vector pF-MGC3
[0180] 1. MGC3 gene amplification and purification The codon-optimized MGC3 gene (SEQ ID NO: 5) was synthesized in Nanjing GenScript Company and cloned into the pUC17 vector to obtain the pUC-MGC3 plasmid vector. The pUC-MGC3 plasmid was used as a template, and MGC3-F and MGC3-R were used as upstream and downstream primers for PCR amplification (the gene sequences of MGC3-F and MGC3-R are shown in SEQ ID NO. 15 and 16). The amplification system is shown in Table 9.
[0181] Table 9 MGC3 gene amplification system
[0182]
[0183] The reaction conditions were: pre-denaturation at 95°C for 5 minutes; denaturation at 94°C for 45 seconds, annealing at 54°C for 45 seconds, extension at 72°C for 1 minute, 35 cycles; extension at 72°C for 10 minutes.
[0184] The PCR product is subjected to gel electrophoresis to verify the size of the target gene, such as Figure 7 As shown, the target band appeared ...
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