3 results about "Thiogalactosides" patented technology

The application discloses a fermentation method for producing thiamine pyrophosphokinase and belongs to the technical field of bioengineering. The BL21 recombinant engineering bacteria containing pET28a-TPK, pET28a-ProS2-TPK or pET28a-SUMO-TPK are constructed, the seed liquid is prepared, the isopropyl-beta-D-thiogalactoside is used for inducing expression, the bacteria are collected, the cells are broken and the supernatant is collected to obtain the TPK crude enzyme liquid. In the fermentation, the LB / TB culture medium containing 50 μg / mL kanamycin or 100 μg / mL ampicillin is used. The SUMO label is introduced to guide the correct folding of the TPK, so that the soluble expression ratio of the TPK is more than 85%, the ratio is improved by 2 times compared with the common system, the inclusion body ratio is greatly reduced, the process is stable and has good repeatability, and an effective scheme is provided for the industrialized large-scale preparation of the TPK.

The preparation method comprises the following steps: (1) preparing a seed culture medium, and culturing at the temperature of 30 + / -1 DEG C until the logarithmic phase to obtain a seed solution; (2) adding the seed solution into a fermentation tank filled with a fermentation basal culture medium, and carrying out two-stage pH regulation and control fermentation; stage 1: under the conditions that the pH is 5.5 + / -0.1, the temperature is 30 + / -0.5 DEG C and the dissolved oxygen DO is 50 + / -5%, carrying out enrichment culture on the thalli until the OD600 is 15-20; stage 2, reducing the pH to 3.8 + / -0.1, reducing the temperature to 28 + / -0.5 DEG C, reducing dissolved oxygen (DO) to 20 + / -5%, and carrying out small molecule peptide synthesis, wherein the reaction time is 18-60 hours; 0.1-0.2 mM isopropyl-beta-D-thiogalactoside is added at the beginning of the stage 2 to continuously induce the expression of the exogenous gene, and the concentration of residual sugar is maintained at 2-5g / L in the synthesis process; and 3) separating and purifying the fermentation broth to obtain the peptide. According to the method, the metabolism of amino acid is enhanced, the degradation of peptide is blocked, the yield of small molecule peptide is increased, and the metabolism has antibacterial and antioxidant effects by virtue of precise fermentation control and an efficient purification process.

The invention belongs to the technical field of biology, and discloses a preparation method of a polyclonal antibody of a gene VII type Newcastle disease virus NP protein, which is characterized by comprising the following steps: step 1, constructing a recombinant plasmid containing a gene VII type Newcastle disease virus NP protein gene, the sequence of the NP protein gene being as shown in SEQ ID NO.1; 2, after the recombinant plasmid is converted into escherichia coli, isopropyl-beta-D-thiogalactoside is added to induce recombinant protein for expressing NP; and 3, purifying the NP recombinant protein, immunizing a New Zealand rabbit to obtain immune serum, and purifying the immune serum to obtain the polyclonal antibody of the Newcastle disease virus NP protein, the antibody can recognize a plurality of linear and conformational epitopes of the Newcastle disease virus NP protein, and the titer is high and can reach 1: 512000. The polyclonal antibody can improve the detection sensitivity and has high tolerance to small antigen variation, and meanwhile, the invention further provides the polyclonal antibody obtained through the preparation method and application of the polyclonal antibody.