Biotechnology Bulletin ›› 2023, Vol. 39 ›› Issue (11): 217-225.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0123
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CHE Yong-mei(
), LIU Guang-chao, GUO Yan-ping, YE Qing, ZHAO Fang-gui, LIU Xin()
Received:2023-02-15
Online:2023-11-26
Published:2023-12-20
Contact:
LIU Xin
E-mail:yongmeiche@163.com;liuxin6080@126.com
CHE Yong-mei, LIU Guang-chao, GUO Yan-ping, YE Qing, ZHAO Fang-gui, LIU Xin. Preparation of Compound Halotolerant Bioinoculant and Study on Its Growth-promoting Effect[J]. Biotechnology Bulletin, 2023, 39(11): 217-225.
| 水平 Level | 葡萄糖 Glucose/（g·L-1） | 酵母膏 Yeast extract/（g·L-1） | NaCl/（g·L-1） |
|---|---|---|---|
| 1 | 8 | 5 | 2.5 |
| 2 | 10 | 10 | 4.5 |
| 3 | 12 | 15 | 6.5 |
Table 1 Factors and levels of orthogonal test
| 水平 Level | 葡萄糖 Glucose/（g·L-1） | 酵母膏 Yeast extract/（g·L-1） | NaCl/（g·L-1） |
|---|---|---|---|
| 1 | 8 | 5 | 2.5 |
| 2 | 10 | 10 | 4.5 |
| 3 | 12 | 15 | 6.5 |
| 因素Factor | 水平Level | ||
|---|---|---|---|
| -1 | 1 | ||
| pH | 5 | 9 | |
| 温度 Temperature/oC | 20 | 30 | |
| 装瓶量 Bottling amount/% | 10 | 30 | |
| 转速 Rotation speed/（r·min-1） | 130 | 180 | |
| 接种量 Inoculation amount/% | 1 | 3 | |
Table 2 Factors and levels of Plackett-Burman design
| 因素Factor | 水平Level | ||
|---|---|---|---|
| -1 | 1 | ||
| pH | 5 | 9 | |
| 温度 Temperature/oC | 20 | 30 | |
| 装瓶量 Bottling amount/% | 10 | 30 | |
| 转速 Rotation speed/（r·min-1） | 130 | 180 | |
| 接种量 Inoculation amount/% | 1 | 3 | |
| 水平 Level | 温度 Temperature/℃ | pH | 转速 Rotation speed/（r·min-1） |
|---|---|---|---|
| α | 33.4 | 10.4 | 200.5 |
| 1 | 30 | 9 | 180 |
| 0 | 25 | 7 | 150 |
| -1 | 20 | 5 | 120 |
| -α | 16.6 | 3.6 | 99.5 |
Table 3 Factors and levels of CCD
| 水平 Level | 温度 Temperature/℃ | pH | 转速 Rotation speed/（r·min-1） |
|---|---|---|---|
| α | 33.4 | 10.4 | 200.5 |
| 1 | 30 | 9 | 180 |
| 0 | 25 | 7 | 150 |
| -1 | 20 | 5 | 120 |
| -α | 16.6 | 3.6 | 99.5 |
Fig. 1 Effects of mixed culture on the abilities of C8 and B4 dissolving potassium, organic and inorganic phosphorus as well as producing auxin The result is the mean ± standard deviation. Different lowercase letters on the column in the figure indicate significant difference between different treatments（P<0.05）, the same below
| 序号 No. | 葡萄糖Glucose（A）/（g·L-1） | 酵母膏Yeast extract（B）/（g·L-1） | NaCl（C）/（g·L-1） | OD600 |
|---|---|---|---|---|
| 1 | 1 | 1 | 1 | 1.17 |
| 2 | 1 | 2 | 2 | 1.28 |
| 3 | 1 | 3 | 3 | 1.12 |
| 4 | 2 | 1 | 2 | 1.30 |
| 5 | 2 | 2 | 3 | 1.35 |
| 6 | 2 | 3 | 1 | 1.32 |
| 7 | 3 | 1 | 3 | 1.34 |
| 8 | 3 | 2 | 1 | 1.27 |
| 9 | 3 | 3 | 2 | 1.32 |
| k1 | 1.27 | 1.28 | 1.28 | |
| k2 | 1.30 | 1.30 | 1.31 | |
| k3 | 1.25 | 1.27 | 1.23 | |
| R | 0.05 | 0.03 | 0.09 |
Table 4 Design and results of orthogonal experiment
| 序号 No. | 葡萄糖Glucose（A）/（g·L-1） | 酵母膏Yeast extract（B）/（g·L-1） | NaCl（C）/（g·L-1） | OD600 |
|---|---|---|---|---|
| 1 | 1 | 1 | 1 | 1.17 |
| 2 | 1 | 2 | 2 | 1.28 |
| 3 | 1 | 3 | 3 | 1.12 |
| 4 | 2 | 1 | 2 | 1.30 |
| 5 | 2 | 2 | 3 | 1.35 |
| 6 | 2 | 3 | 1 | 1.32 |
| 7 | 3 | 1 | 3 | 1.34 |
| 8 | 3 | 2 | 1 | 1.27 |
| 9 | 3 | 3 | 2 | 1.32 |
| k1 | 1.27 | 1.28 | 1.28 | |
| k2 | 1.30 | 1.30 | 1.31 | |
| k3 | 1.25 | 1.27 | 1.23 | |
| R | 0.05 | 0.03 | 0.09 |
| 序号 No. | 因素Factor | OD600 | ||||
|---|---|---|---|---|---|---|
| pH | 温度Temperature/℃ | 装瓶量Bottling volunm/% | 转速Rotating speed/（r·min-1） | 接种量Inoculation amount/% | ||
| 1 | -1 | 1 | 1 | 1 | -1 | 0.79 |
| 2 | 1 | -1 | 1 | 1 | -1 | 0.81 |
| 3 | -1 | 1 | -1 | 1 | 1 | 1.12 |
| 4 | -1 | 1 | 1 | -1 | 1 | 1.29 |
| 5 | 1 | -1 | -1 | -1 | 1 | 1.37 |
| 6 | -1 | -1 | -1 | -1 | -1 | 1.14 |
| 7 | -1 | -1 | -1 | 1 | -1 | 0.80 |
| 8 | -1 | -1 | 1 | -1 | 1 | 1.11 |
| 9 | 1 | 1 | -1 | 1 | 1 | 1.30 |
| 10 | 1 | -1 | 1 | 1 | 1 | 1.48 |
| 11 | 1 | 1 | 1 | -1 | -1 | 1.20 |
| 12 | 1 | 1 | -1 | -1 | -1 | 1.10 |
Table 5 Plackett-Burman design and results
| 序号 No. | 因素Factor | OD600 | ||||
|---|---|---|---|---|---|---|
| pH | 温度Temperature/℃ | 装瓶量Bottling volunm/% | 转速Rotating speed/（r·min-1） | 接种量Inoculation amount/% | ||
| 1 | -1 | 1 | 1 | 1 | -1 | 0.79 |
| 2 | 1 | -1 | 1 | 1 | -1 | 0.81 |
| 3 | -1 | 1 | -1 | 1 | 1 | 1.12 |
| 4 | -1 | 1 | 1 | -1 | 1 | 1.29 |
| 5 | 1 | -1 | -1 | -1 | 1 | 1.37 |
| 6 | -1 | -1 | -1 | -1 | -1 | 1.14 |
| 7 | -1 | -1 | -1 | 1 | -1 | 0.80 |
| 8 | -1 | -1 | 1 | -1 | 1 | 1.11 |
| 9 | 1 | 1 | -1 | 1 | 1 | 1.30 |
| 10 | 1 | -1 | 1 | 1 | 1 | 1.48 |
| 11 | 1 | 1 | 1 | -1 | -1 | 1.20 |
| 12 | 1 | 1 | -1 | -1 | -1 | 1.10 |
| 运行序 Sequence | 温度 Temperature/℃ | pH | 转速Rotational speed/（r·min-1） | OD600 |
|---|---|---|---|---|
| 1 | 1 | -1 | 1 | 1.035 |
| 2 | -1 | -1 | -1 | 1.028 |
| 3 | 1 | 1 | 1 | 1.173 |
| 4 | 0 | 0 | 1 | 1.320 |
| 5 | 33.409 | 0 | 1 | 1.241 |
| 6 | 0 | 0 | 99.546 | 1.273 |
| 7 | 0 | 0 | 1 | 1.350 |
| 8 | 0 | 0 | 200.454 | 0.984 |
| 9 | -1 | 1 | -1 | 1.126 |
| 10 | -1 | 1 | 1 | 1.011 |
| 11 | 0 | 0 | 1 | 1.363 |
| 12 | 0 | 0 | 1 | 1.285 |
| 13 | 0 | 10.364 | 1 | 1.118 |
| 14 | 1 | 1 | -1 | 1.329 |
| 15 | 1 | -1 | -1 | 1.285 |
| 16 | 16.591 | 0 | 1 | 0.925 |
| 17 | 0 | 3.636 | 1 | 1.044 |
| 18 | 0 | 0 | 1 | 1.387 |
| 19 | -1 | -1 | 1 | 0.998 |
| 20 | 0 | 0 | 1 | 1.387 |
Table 6 Results of CCD design
| 运行序 Sequence | 温度 Temperature/℃ | pH | 转速Rotational speed/（r·min-1） | OD600 |
|---|---|---|---|---|
| 1 | 1 | -1 | 1 | 1.035 |
| 2 | -1 | -1 | -1 | 1.028 |
| 3 | 1 | 1 | 1 | 1.173 |
| 4 | 0 | 0 | 1 | 1.320 |
| 5 | 33.409 | 0 | 1 | 1.241 |
| 6 | 0 | 0 | 99.546 | 1.273 |
| 7 | 0 | 0 | 1 | 1.350 |
| 8 | 0 | 0 | 200.454 | 0.984 |
| 9 | -1 | 1 | -1 | 1.126 |
| 10 | -1 | 1 | 1 | 1.011 |
| 11 | 0 | 0 | 1 | 1.363 |
| 12 | 0 | 0 | 1 | 1.285 |
| 13 | 0 | 10.364 | 1 | 1.118 |
| 14 | 1 | 1 | -1 | 1.329 |
| 15 | 1 | -1 | -1 | 1.285 |
| 16 | 16.591 | 0 | 1 | 0.925 |
| 17 | 0 | 3.636 | 1 | 1.044 |
| 18 | 0 | 0 | 1 | 1.387 |
| 19 | -1 | -1 | 1 | 0.998 |
| 20 | 0 | 0 | 1 | 1.387 |
| 来源Source | 自由度Degree of freedom | Seq SS | Adj SS | Adj MS | F | P |
|---|---|---|---|---|---|---|
| 回归 | 9 | 0.439 222 | 0.439 222 | 0.048 802 | 25.01 | 0.000 |
| 线性 | 3 | 0.195 425 | 0.123 644 | 0.041 215 | 21.12 | 0.000 |
| 平方 | 3 | 0.234 599 | 0.234 599 | 0.078 200 | 40.07 | 0.000 |
| 交互作用 | 3 | 0.009 197 | 0.009 197 | 0.003 066 | 1.57 | 0.257 |
| 残差误差 | 10 | 0.019 517 | 0.019 517 | 0.001 952 | ||
| 失拟 | 5 | 0.011 442 | 0.011 442 | 0.002 288 | 1.42 | 0.356 |
| 纯误差 | 5 | 0.008 075 | 0.008 075 | 0.001 615 | ||
| 合计 | 19 | 0.458 739 |
Table 7 Variance analysis of CCD design
| 来源Source | 自由度Degree of freedom | Seq SS | Adj SS | Adj MS | F | P |
|---|---|---|---|---|---|---|
| 回归 | 9 | 0.439 222 | 0.439 222 | 0.048 802 | 25.01 | 0.000 |
| 线性 | 3 | 0.195 425 | 0.123 644 | 0.041 215 | 21.12 | 0.000 |
| 平方 | 3 | 0.234 599 | 0.234 599 | 0.078 200 | 40.07 | 0.000 |
| 交互作用 | 3 | 0.009 197 | 0.009 197 | 0.003 066 | 1.57 | 0.257 |
| 残差误差 | 10 | 0.019 517 | 0.019 517 | 0.001 952 | ||
| 失拟 | 5 | 0.011 442 | 0.011 442 | 0.002 288 | 1.42 | 0.356 |
| 纯误差 | 5 | 0.008 075 | 0.008 075 | 0.001 615 | ||
| 合计 | 19 | 0.458 739 |
Fig. 2 Response surface of the interactive effect of different factors on the biomass of bacteria strain A: Effect of rotating speed and pH on the biomass of bacteria strain. B: Effect of rotating speed and temperature on the biomass of bacteria strain. C: Effect of temperature and pH on the biomass of bacteria strain
Fig. 3 Effect of fermentation optimization on biomass, function of dissolving potassium, inorganic phosphorus and organic phosphorus as well as producing auxinof mixed strains of C8 and B4
Fig. 4 Effect of compound bioinoculant of C8 and B4 on the growth of aboveground and underground parts of tobacco under salt stress
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