A kind of dual inactivated vaccine of mycoplasma pneumonia and chlamydia psittaci and preparation method thereof
A dual inactivated vaccine, the technology of Chlamydia psittaci, applied in the direction of Chlamydia antigen components, microorganism-based methods, biochemical equipment and methods, etc. Safety, the effect of important application value
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Embodiment 1
[0069] Embodiment 1, the seed batch preparation of mycoplasma mycoplasma goat subspecies and mycoplasma ovis pneumoniae
[0070] The inventor of the present invention is located in my country's Chongqing City, Anhui Province, Qinghai Province, Yunnan Province, Xinjiang Uygur Autonomous Region, Guizhou Province, Guangxi Zhuang Autonomous Region, and Hohhot, Keshigten Banner, Ordos, Siziwang Banner, Ewenki Banner, and Lanzhou in the Inner Mongolia Autonomous Region. The sheep farms in Qi and Wuyuan counties collected 2684 samples of lung tissue and throat swabs from sheep with suspected mycoplasma pneumonia. After separation and testing, the dominant epidemic strains of Mycoplasma mycoplasma goat and Mycoplasma ovis pneumoniae were determined. strain. Specific steps are as follows:
[0071] 1. Primary screening
[0072] (1) Put the sample into a culture bottle filled with mycoplasma liquid medium (throat swabs are put in directly, sheep lung tissue is put in after grinding), and...
Embodiment 2
[0089] Embodiment 2, the screening determination of inactivated bacterium vaccine strain and bacterial strain for inspection
[0090] 1. Morphological observation
[0091] A. Morphological observation of Mycoplasma mycoplasma goat subspecies
[0092] 1. Select 5 strains from the mycoplasma goat subspecies screened in step 3 in Example 1, named successively as GT-01 strain, GT-02 strain, GT-03 strain, GT-04 strain and GT-05 strain , respectively from Wuyuan County, Inner Mongolia, Chongqing, Guizhou, Xinjiang Uygur Autonomous Region and Ordos, Inner Mongolia.
[0093] 2. Carry out Dienes staining to mycoplasma goat subspecies (GT-01 strain, GT-02 strain, GT-03 strain, GT-04 strain or GT-05 strain), and then place it under a microscope (200×) to observe the colony form.
[0094] See the experimental results figure 1 (A-E are GT-01 strain, GT-02 strain, GT-03 strain, GT-04 strain and GT-05 strain). The results showed that the colonies of the five strains of Mycoplasma goat s...
Embodiment 3
[0170] Embodiment 3, the expression of MOMP protein (the main outer membrane protein of Chlamydia psittaci)
[0171] 1. Construction of recombinant plasmid pET-22b-MOMP
[0172] 1. Because the MOMP gene of Chlamydia psittaci is highly conserved among the same species, the MOMP gene (GenBank: L25436.1), a typical representative of Cps, was selected for codon optimization. The nucleotide sequence shown in Sequence 1 in the sequence listing after codon optimization (ie MOMP gene) was artificially synthesized.
[0173] 2. Use the nucleotide sequence of the MOMP gene synthesized in step 1 as a template, and use 5'-TATT CATATG TTGCCTGTAGGGAACCCAGCT-3' (the underline is the recognition site of restriction endonuclease NdeI) and 5'-GCGC CTCGAG The primer pair consisting of CTAGAATCTGAATTGAGCATT-3' (the underline is the recognition site of restriction endonuclease XhoI) was subjected to PCR amplification to obtain a PCR amplification product of about 1130bp.
[0174] 3. Take the P...
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