Yeast with high yield of acid-stage cellulose and diastatisch
A technology of acid cellulase and acid amylase, applied in the field of microorganisms, can solve the problems of little known research on acid yeast
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Embodiment 1
[0017] Example 1 strain isolation
[0018] Enrichment medium (g / L): (NH 4 ) 2 SO 4 3.0, KCl0.1, K 2 HPO 4 0.5, MgSO 4 ·7H 2 O0.5, Ca(NO 3 ) 2 0.01, FeSO 4 ·7H 2 O3.0, glucose 2, pH 2.0-2.5.
[0019] Separation medium (g / L): (NH 4 ) 2 SO 4 2.0, KCl0.1, MgSO 4 ·7H 2 O0.25, Ca(NO 3 ) 2 0.01, glucose 1.0, yeast extract 0.1, pH 2.5-3.0.
[0020] Potato liquid medium: 200g potato juice, 20g glucose, pH 3.0.
[0021] Potato solid medium: 200g potato, 20g glucose, 25g agar, pH3.0.
[0022] Take 10mL of the tail liquid of uranium leaching microorganisms in Jiangxi uranium mines, add it to 100mL enrichment medium for 72 hours at 30°C and 160rpm shaking culture, pick single colonies and transfer them to the separation medium and culture them for two generations under the same conditions, repeat in parallel 3 times. The enriched culture was streaked and diluted in potato solid medium to obtain a single colony, and the streaked dilution was repeated several times to ob...
Embodiment 2
[0024] Embodiment 2 bacterial strain liquid fermentation produces enzyme analysis
[0025] Cellulase fermentation medium (g / L): (NH 4 ) 2 SO 4 2.0,K 2 HPO 4 1.0,; MgSO 4 ·7H 2 O0.5; wheat bran 10g, pH3.0.
[0026] Amylase fermentation medium (g / L): (NH 4 ) 2 SO 4 2.0,K 2 HPO 4 1.0,; MgSO 4 ·7H 2 O0.5; Potato starch 10g, pH3.0.
[0027] DNS method: The specific method is as follows: at pH 3.0, 50°C, 1 mL of reaction system includes 100 μL of appropriate diluted enzyme solution, 900 μL of substrate (sodium carboxymethylcellulose or soluble starch), react for 10 minutes, add 1.5 mL DNS terminates the reaction and boils for 5 minutes. After cooling to room temperature, the OD value was measured at 540 nm. One enzyme activity unit (U) is defined as the amount of enzyme that releases 1 μmol of reducing sugar per minute under given conditions.
[0028] pNPG method: the substrate p-nitrophenyl β-D glucoside (pNPG) was dissolved in 250uL of pH4.0 buffer at a concentrat...
Embodiment 3
[0030] The optimum pH of different enzymes of embodiment 3
[0031] Separate and purify β-glucosidase, cellulase and amylase, and carry out enzymatic reaction at different pH to determine their optimum pH. The substrates were tested for enzyme activity in 0.1mol / L citric acid-disodium hydrogen phosphate buffer solution with different pH at 50°C. The results showed that the optimum pH of β-glucosidase was 3.0, and there was more than 70% relative enzyme activity at pH 2.5~6.0 ( figure 1 ). The optimal pH of cellulase is 3.5, and there is more than 60% relative enzyme activity at pH 3.0~5.0 ( figure 2 ). The optimal pH of amylase is 4.5, and there is more than 50% relative enzyme activity at pH 3.5~5.5 ( image 3 ). The above results indicated that the optimal pH of the glycoside hydrolase produced by strain RBS-9 was in the acidic range, and it had higher activity under acidic and neutral pH conditions.
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