Method for continuously producing xylanase by using clostridium
A technology of xylanase and Clostridium, which is applied in the field of genetic engineering, can solve the problems of ignoring proteins, etc., and achieve the effects of high enzyme activity, continuous enzyme production, and easy separation
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Embodiment 1
[0046] Embodiment 1: Carrier selection of continuous culture
[0047] (1) Carrier material: The carrier refers to the relevant fermentation papers, and the specific names are shown in Table 1, corresponding to figure 1 .
[0048] Table 1 Carrier material and number
[0049]
[0050] Remarks: Polyethylene (small) is a round polyethylene with a diameter of 1.3cm; polyethylene (large) is a round polyethylene with a diameter of 2.4cm; the diameter of the resin (porous) is 2.5cm, and the hole is uneven, 0.3- 0.5 cm; the diameter of the clay material is 2.2 cm; the resin is polyethylene resin; the sponge analogue (macropore) is honeycomb-shaped pores of 0.2-0.5 cm.
[0051] (2) Immobilized culture:
[0052] (i) all carrier materials are cut into appropriate sizes, and sterilized by high temperature and high pressure for standby;
[0053] (ii) Transfer the Clostridium acetobutylicum B3 bacterium sludge after plate activation to P2 seed medium (containing 20 μg / L thiamphenicol ...
Embodiment 2
[0057] Embodiment 2: Cotton towel carrier continuous culture
[0058] On the basis of Example 1, only the cotton fiber towel (5) was selected as the carrier for surface-immobilized continuous fermentation, and the experiment was expanded to a 3L fermenter experiment. In addition, in step (iii), the time for sampling and liquid replacement was changed to 12 hours, and the culture time was changed to one week.
[0059] The dry weight of each batch of fermented cotton towel carriers was weighed respectively, and the changes in dry weight are shown in Table 2. In addition, the bacterial cell attachment of cotton towels during the cultivation process can be found in image 3 . The above results indicated that the cotton towel carrier was beneficial to the formation of biofilm.
[0060] Table 2 Changes in dry weight of biofilms
[0061]
Embodiment 3
[0062] Example 3: Determination of xylanase activity——DNS method.
[0063] (1) Add 25 μL of enzyme solution diluted to a certain concentration, 500 μL of xylan solution with a concentration of 5 mg / mL and 225 μL of phosphate buffer solution into each 10 mL centrifuge tube;
[0064] (2) After the system was reacted in a constant temperature water bath at 65°C for 15 minutes, it was immediately placed on ice and 1 mL of prepared DNS solution was added to each reaction system;
[0065] (3) After reacting in a boiling water bath for 5 minutes, immediately place it in an ice water bath, let it cool to room temperature, and add pure water to make it to 5 mL;
[0066] (4) Measure its absorbance at 540 nm by using a UV spectrophotometer.
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