GB/T 38569-2020Performance evaluation of industrial microorganism strain—Raman spectroscopy (English PDF)

3.1 Performance evaluation of industrial microorganism strain

Use Raman spectroscopy to carry out performance consistency evaluation of industrial microorganism strain.

3.2 Reference strain

The identified stock strain, whose fermentation performance is clear, for fermentation production.

7.3 Sample preparation

7.3.1 Respectively transfer 1 mL of the cultured reference strain and the to-be-evaluated strain into a sterile tube.

7.3.2 Select the corresponding speed for centrifugation; discard the supernatant; add 1 mL of sterile double-distilled water; mix the bacteria.

7.3.3 Repeat 2 times in accordance with 7.3.2; transfer 10 µL of bacterial solution to a sterile tube; add 990 µL of sterile double-distilled water; mix well; pipette 2 µL of diluted bacterial solution; place it in a clean calcium fluoride glass slide for sample application; air-dry at room temperature in a sterile environment.

7.4 Measurement

7.4.1 Place the air-dried calcium fluoride glass slide on the micro Raman spectrometer stage; select a single cell in the transmissive light field mode; switch to the spectrum acquisition mode; measure the Raman spectrum signal.

7.4.2 Select at least 20 single cells randomly from each sample as the sample signal.

7.4.3 In the sample application area, collect 3 acellular signals; calculate the average value as the background signal.

7.5 Data processing

7.5.1 Perform background signal sub traction (sample signal minus background

signal), baseline correction ( >5th order function) and area-based normalization of the raw data of single-cell Raman spectrum.

7.5.2

Calculate the signal-to-noise ratio, where the signal uses the signal intensity corresponding to the sharp Raman peak at 1 001 cm-1; discard data whose signal-to-noise ratio is less than 3; supplement the number of data, so that the to-be-evaluated strain group and the reference strain group have the same number of data. […]