Rhodopseudomonas palustris Atps2 protein, preparation method and application thereof
A swamp red pseudomonas, protein technology, applied in the direction of botanical equipment and methods, biochemical equipment and methods, applications, etc., to achieve the effects of short culture time, environmental friendliness, and inhibition of pathogenicity
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Embodiment 1
[0035] Preparation method of Rhodopseudomonas palustris Atps2 protein,
[0036] Include the following steps:
[0037] (1) Prokaryotic expression of Rhodopseudomonas palustris ATPS2 gene in Escherichia coli
[0038] Constructing the vector: connecting the nucleotide sequence of the ATPS2 gene of Rhodopseudomonas palustris to the PET32a plasmid containing the ampicillin resistance gene;
[0039] Escherichia coli transformation: the plasmid constructed in the previous step is introduced into the expression competent cell BL21, spread on the LB plate containing ampicillin resistance and cultivated; the medium of the LB plate containing ampicillin resistance, the formula For: yeast extract 5g / L, peptone 10g / L, NaCl 5gm, ampicillin 100mg / L, agar 15g / L, pH=7.0;
[0040] Expansion of transformed strains and protein induction: transfer the single colony cultivated in the previous step into a 120mL Erlenmeyer flask and culture at 37°C until the stable phase; then inoculate the bacteri...
Embodiment 2
[0044] Application of Atps2 Protein from Rhodopseudomonas palustris in Inhibiting Magnaporthe grisea
[0045] (1) Application of Atps2 protein from Rhodopseudomonas palustris in inhibiting the formation of conidia of blast fungus oryzae
[0046] Select the Rhodopseudomonas palustris Atps2 protein (final concentration is respectively 0 μ g / ml, 50 μ g / ml, 100 μ g / ml, 200 μ g / ml, 500 μ g / ml) prepared in embodiment 1, it is added to blast fungus meristem Spore Liquid (1×10 5 pcs / ml) for mixing, then 30 μl was dropped onto the hydrophobic membrane, and each group (corresponding to CK group, 50 μg / ml group, 100 μg / ml group, 200 μg / ml group, 500 μg / ml group) set 3 Repeat, observe microscopically after 8 hours and count the appressor formation rate, and evaluate the respective inhibitory effects;
[0047] Such as figure 2 As shown, the results show that the final concentration of Rhodopseudomonas palustris Atps2 protein treatment with a final concentration of 100ug / mL has a signif...
Embodiment 3
[0052] Referring to Example 2 "Application of the Rhodopseudomonas palustris Atps2 Protein in Inhibiting the Formation of Pyricularia Oryzae Conidia Appresses", after prokaryotic expression, the concentration of 500ug / mL 3-hydroxyacyl dehydratase FabA, The outer membrane protein assembly factor BamA and Atps2 protein were tested, and the appressor formation rate was counted. Among them, the 3-hydroxyacyl dehydratase FabA is derived from 3-hydroxyacyl-[acyl chain protein] dehydratase, the base pair is 525bp, and the protein molecular weight is 18.8kD; the outer membrane protein assembly factor BamA is derived from the outer membrane protein assembly factor, base The base pair is 2454bp, and the protein molecular weight is 93.5kD.
[0053] Such as Figure 4 As shown, the formation rate of conidia of Magnaporthe grisea after treatment with Atps2 protein was less than 10%, which was significantly lower than that of CK control (** represents a very significant difference), while t...
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