Ester-producing yeast strain and application thereof
A Pichia pastoris and strain technology, applied in the field of microorganisms, can solve the problems of poor substrate adaptability, low ester production, poor environmental tolerance, etc., and achieve the effects of improving product quality, improving sensory flavor, and enriching product taste
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Embodiment 1
[0046] Isolation and identification of yeast strains in embodiment 1 of the present invention
[0047] Dissolve the cellar mud sample of the Baogu Distillery in Enshi, Hubei, in sterile water and mix evenly, draw the bacterial suspension and dilute it 10 times serially to prepare 10 -5 、10 -6Bacterial suspension, spread on YPD medium, culture at 30°C for 24-48h, make slices and observe the morphology of yeast under a microscope, and observe the characteristics of single colonies on the plate at the same time, isolate and purify the strains with typical yeast colony characteristics, and inoculate Store in YPD slant medium at 4°C. A strain of bacteria was obtained, the colony texture of the bacteria was cheese-like, the color was milky white, and the surface was smooth; the microscopic shape was oval, the size was 3.0×7.5μm, and the microscopic observation showed budding. The genome of the yeast strain was extracted, using NL1 (5'-GCATATCAATAAGCGGAGGAAAAG-3') and NL4 (5'-GGTCC...
Embodiment 2
[0051] Example 2 Analysis of Substrate Utilization Characteristics of Pichia pastoris C4.12 Strain
[0052] Inoculate the C4.12 strain of Pichia wisterii into a test tube containing 5mL of LYPD liquid medium, culture at 30°C and 180rpm for 20h, and then inoculate it into 300uL of carbon source liquid medium according to the inoculation amount of 3%. In the 100-well culture plate, prepare the Bioscreen instrument for measurement on the machine, set the parameters: temperature 30°C, time 24h, wavelength 600nm, measure the data every 30min, and then measure the growth curve, with OD600 nm A growth rate of more than 50% is considered growth.
[0053]
[0054] OD1—OD600nm value at the beginning;
[0055] OD2—OD600nm value at the end;
[0056] Such as image 3 As shown, the results show that the C4.12 strain of Pichia wisteria can utilize the above-mentioned 15 carbon sources, and when utilizing glucose and raffinose, it preferentially reaches the logarithmic phase and the sta...
Embodiment 3
[0057] Example 3 Tolerance Analysis of Pichia pastoris C4.12 strain
[0058] (1) pH tolerance test: Inoculate Pichia kudriazzia C4.12 strain into a test tube containing 5mL YPD liquid medium, culture at 30°C and 180rpm for 20h, and then use 3% inoculation amount Inoculate into a 100-well culture plate with 300uL YPD liquid medium, adjust the pH of YPD liquid medium to 2.0, 3.0, 4.0, 5.0, 6.0 respectively, prepare the Bioscreen instrument for measurement, set parameters: temperature 30°C, time 24h , the wavelength is 600nm, the data is measured every 30min, and then the growth curve is determined.
[0059] Such as Figure 4 As shown, the growth curves of the Pichia wisteria C4.12 strain at pH 3.0, 4.0, 5.0, and 6.0 showed a typical S-shaped curve model, and the growth curve of the strain at pH 2.0 It is approximately a straight line, that is, the strain can grow normally under the conditions of pH 3.0, 4.0, 5.0, and 6.0, but the strain cannot grow normally under the condition...
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