Chinese Bulletin of Botany ›› 2023, Vol. 58 ›› Issue (6): 917-925.DOI: 10.11983/CBB22228
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Xiaohui Feng1,2,3, Xuetong Yan2,3, Keyuan Zheng2,3, Qiang Zhou1, Weizhong Zhang4, Quanyong Wang4, Mulan Zhu2,3,*()
Received:
2022-09-29
Accepted:
2023-02-28
Online:
2023-11-01
Published:
2023-11-27
Contact:
* E-mail: mlzhu@cemps.ac.cn
Xiaohui Feng, Xuetong Yan, Keyuan Zheng, Qiang Zhou, Weizhong Zhang, Quanyong Wang, Mulan Zhu. In vitro Culture of Taxus Rich in Taxanes[J]. Chinese Bulletin of Botany, 2023, 58(6): 917-925.
Treatments | 75% ethanol (s) | Disinfectant (min) | Pollution rate (%) | Survival rate (%) |
---|---|---|---|---|
1 | 10 | 3 | 68.9±5.09a | 31.1±5.09f |
2 | 10 | 5 | 35.6±3.85c | 57.8±3.85bcd |
3 | 10 | 7 | 20.0±3.33de | 62.2±3.85bc |
4 | 30 | 3 | 54.4±5.09b | 45.6±5.09e |
5 | 30 | 5 | 25.6±5.09d | 74.4±5.09a |
6 | 30 | 7 | 12.2±1.92ef | 64.4±8.38b |
7 | 50 | 3 | 41.1±5.09c | 51.1±1.92de |
8 | 50 | 5 | 16.7±3.33e | 64.4±5.09b |
9 | 50 | 7 | 5.6±1.92f | 53.3±5.77de |
Table 1 Effects of disinfection treatments on the contamination rate and survival rate of explants
Treatments | 75% ethanol (s) | Disinfectant (min) | Pollution rate (%) | Survival rate (%) |
---|---|---|---|---|
1 | 10 | 3 | 68.9±5.09a | 31.1±5.09f |
2 | 10 | 5 | 35.6±3.85c | 57.8±3.85bcd |
3 | 10 | 7 | 20.0±3.33de | 62.2±3.85bc |
4 | 30 | 3 | 54.4±5.09b | 45.6±5.09e |
5 | 30 | 5 | 25.6±5.09d | 74.4±5.09a |
6 | 30 | 7 | 12.2±1.92ef | 64.4±8.38b |
7 | 50 | 3 | 41.1±5.09c | 51.1±1.92de |
8 | 50 | 5 | 16.7±3.33e | 64.4±5.09b |
9 | 50 | 7 | 5.6±1.92f | 53.3±5.77de |
Treatments | NAA (mg·L-1) | 6-BA (mg·L-1) | Proline (g·L-1) | Expansion rate (%) |
---|---|---|---|---|
1 | 0.1 | 1 | 0 | 28.9±5.09g |
2 | 0.1 | 2 | 0.1 | 53.3±8.82ef |
3 | 0.1 | 3 | 0.3 | 62.3±2.02de |
4 | 0.2 | 1 | 0.5 | 58.9±5.09ef |
5 | 0.2 | 2 | 0.7 | 76.7±6.67abc |
6 | 0.2 | 3 | 1 | 86.7±5.77ab |
7 | 0.3 | 1 | 0 | 47.8±6.93f |
8 | 0.3 | 2 | 0.1 | 51.1±15.40ef |
9 | 0.3 | 3 | 0.3 | 86.7±5.77ab |
10 | 0.4 | 1 | 0.5 | 55.6±5.09ef |
11 | 0.4 | 2 | 0.7 | 90.0±6.67a |
12 | 0.4 | 3 | 1 | 63.3±3.33cde |
13 | 0.5 | 1 | 0 | 52.2±1.92ef |
14 | 0.5 | 2 | 0.1 | 75.6±8.39bcd |
15 | 0.5 | 3 | 0.3 | 81.1±10.18ab |
16 | 0.6 | 1 | 0.5 | 64.4±11.71cde |
17 | 0.6 | 2 | 0.7 | 57.8±6.94ef |
18 | 0.6 | 3 | 1 | 46.7±5.77f |
Table 2 Effect of different additive combinations on the bottom absorbent surface
Treatments | NAA (mg·L-1) | 6-BA (mg·L-1) | Proline (g·L-1) | Expansion rate (%) |
---|---|---|---|---|
1 | 0.1 | 1 | 0 | 28.9±5.09g |
2 | 0.1 | 2 | 0.1 | 53.3±8.82ef |
3 | 0.1 | 3 | 0.3 | 62.3±2.02de |
4 | 0.2 | 1 | 0.5 | 58.9±5.09ef |
5 | 0.2 | 2 | 0.7 | 76.7±6.67abc |
6 | 0.2 | 3 | 1 | 86.7±5.77ab |
7 | 0.3 | 1 | 0 | 47.8±6.93f |
8 | 0.3 | 2 | 0.1 | 51.1±15.40ef |
9 | 0.3 | 3 | 0.3 | 86.7±5.77ab |
10 | 0.4 | 1 | 0.5 | 55.6±5.09ef |
11 | 0.4 | 2 | 0.7 | 90.0±6.67a |
12 | 0.4 | 3 | 1 | 63.3±3.33cde |
13 | 0.5 | 1 | 0 | 52.2±1.92ef |
14 | 0.5 | 2 | 0.1 | 75.6±8.39bcd |
15 | 0.5 | 3 | 0.3 | 81.1±10.18ab |
16 | 0.6 | 1 | 0.5 | 64.4±11.71cde |
17 | 0.6 | 2 | 0.7 | 57.8±6.94ef |
18 | 0.6 | 3 | 1 | 46.7±5.77f |
Figure 1 Tissue culture of Taxus (A) Image of bottom absorbent surface expands from the front (bar=10 mm); (B) Image of bottom absorbent surface expands from the underside (bar=5 mm); (C) Axillary bud induction (bar=10 mm); (D) The original basis of adventitious bud induction (bar=10 mm); (E) High-frequency synchronous growth (bar=10 mm); (F) Adventitious bud elongation (bar=10 mm); (G) Biomass amplification (bar=20 mm)
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Axillary buds induction rate (%) |
---|---|---|---|
1 | 0.5 | 0.05 | 63.3±3.33ef |
2 | 0.5 | 0.1 | 48.9±5.09g |
3 | 1 | 0.05 | 81.1±1.92ab |
4 | 1 | 0.1 | 88.9±5.09a |
5 | 1 | 0.2 | 68.9±5.09cde |
6 | 1 | 0.3 | 64.4±5.09ef |
7 | 2 | 0.05 | 73.3±3.34bcd |
8 | 2 | 0.1 | 75.6±3.85bc |
9 | 2 | 0.2 | 65.6±5.09def |
10 | 2 | 0.3 | 58.9±5.09f |
11 | 3 | 0.1 | 60.0±3.33f |
12 | 3 | 0.2 | 43.3±3.34g |
Table 3 Effect of different concentrations of 6-BA and NAA on axillary bud induction
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Axillary buds induction rate (%) |
---|---|---|---|
1 | 0.5 | 0.05 | 63.3±3.33ef |
2 | 0.5 | 0.1 | 48.9±5.09g |
3 | 1 | 0.05 | 81.1±1.92ab |
4 | 1 | 0.1 | 88.9±5.09a |
5 | 1 | 0.2 | 68.9±5.09cde |
6 | 1 | 0.3 | 64.4±5.09ef |
7 | 2 | 0.05 | 73.3±3.34bcd |
8 | 2 | 0.1 | 75.6±3.85bc |
9 | 2 | 0.2 | 65.6±5.09def |
10 | 2 | 0.3 | 58.9±5.09f |
11 | 3 | 0.1 | 60.0±3.33f |
12 | 3 | 0.2 | 43.3±3.34g |
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Adventitious buds induction rate (%) |
---|---|---|---|
1 | 0.2 | 0.02 | 32.2±5.09e |
2 | 0.4 | 0.04 | 47.8±3.85d |
3 | 0.6 | 0.06 | 62.2±1.92bc |
4 | 0.8 | 0.08 | 68.9±3.84b |
5 | 1.0 | 0.10 | 77.8±1.92a |
6 | 1.2 | 0.12 | 67.8±5.09bc |
7 | 1.4 | 0.14 | 61.1±3.84c |
Table 4 Effect of different concentration of 6-BA and NAA on the original basis of adventitious bud induction
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Adventitious buds induction rate (%) |
---|---|---|---|
1 | 0.2 | 0.02 | 32.2±5.09e |
2 | 0.4 | 0.04 | 47.8±3.85d |
3 | 0.6 | 0.06 | 62.2±1.92bc |
4 | 0.8 | 0.08 | 68.9±3.84b |
5 | 1.0 | 0.10 | 77.8±1.92a |
6 | 1.2 | 0.12 | 67.8±5.09bc |
7 | 1.4 | 0.14 | 61.1±3.84c |
Treatments | NAA (mg·L-1) | 6-BA (mg·L-1) | High-frequency synchronous growth rate (%) | Average number of adventitious buds |
---|---|---|---|---|
1 | 0.05 | 0.25 | 38.9±5.09e | 12.2±2.65c |
2 | 0.05 | 0.5 | 55.6±5.09cd | 14.2±2.78bc |
3 | 0.1 | 0.5 | 50.0±6.67d | 20.1±3.66b |
4 | 0.1 | 1 | 82.2±1.92a | 31.5±2.93a |
5 | 0.2 | 1 | 72.2±3.85b | 33.4±3.67a |
6 | 0.2 | 2 | 63.3±3.34bc | 29.7±2.94a |
Table 5 Effect of different medium formulations on the original basis of adventitious bud high-frequency synchronous growth
Treatments | NAA (mg·L-1) | 6-BA (mg·L-1) | High-frequency synchronous growth rate (%) | Average number of adventitious buds |
---|---|---|---|---|
1 | 0.05 | 0.25 | 38.9±5.09e | 12.2±2.65c |
2 | 0.05 | 0.5 | 55.6±5.09cd | 14.2±2.78bc |
3 | 0.1 | 0.5 | 50.0±6.67d | 20.1±3.66b |
4 | 0.1 | 1 | 82.2±1.92a | 31.5±2.93a |
5 | 0.2 | 1 | 72.2±3.85b | 33.4±3.67a |
6 | 0.2 | 2 | 63.3±3.34bc | 29.7±2.94a |
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Adventitious bud elongation rate (%) | Average seedling height (mm) |
---|---|---|---|---|
1 | 0.1 | 0.01 | 35.5±6.94c | 17.3±0.85d |
2 | 0.3 | 0.03 | 40.0±3.33c | 22.7±1.34c |
3 | 0.5 | 0.05 | 74.4±5.09b | 28.4±0.94b |
4 | 0.7 | 0.07 | 92.2±1.92a | 33.4±1.56a |
5 | 1 | 0.1 | 80.0±5.77b | 29.9±0.69ab |
Table 6 Effect of medium formulations on the elongation of adventitious buds
Treatments | 6-BA (mg·L-1) | NAA (mg·L-1) | Adventitious bud elongation rate (%) | Average seedling height (mm) |
---|---|---|---|---|
1 | 0.1 | 0.01 | 35.5±6.94c | 17.3±0.85d |
2 | 0.3 | 0.03 | 40.0±3.33c | 22.7±1.34c |
3 | 0.5 | 0.05 | 74.4±5.09b | 28.4±0.94b |
4 | 0.7 | 0.07 | 92.2±1.92a | 33.4±1.56a |
5 | 1 | 0.1 | 80.0±5.77b | 29.9±0.69ab |
Treatments | PG (mg·L-1) | Average stem thickness (mm) | Average seedling biomass (mg) |
---|---|---|---|
1 | 0 | 0.6±0.05c | 283±15d |
2 | 20 | 0.8±0.10b | 483±12b |
3 | 50 | 1.1±0.15a | 542±13a |
4 | 80 | 0.9±0.06ab | 490±15b |
5 | 100 | 0.8±0.15bc | 353±20c |
Table 7 Effects of different concentrations of phloroglucinol (PG) on Taxus biomass amplification phase I
Treatments | PG (mg·L-1) | Average stem thickness (mm) | Average seedling biomass (mg) |
---|---|---|---|
1 | 0 | 0.6±0.05c | 283±15d |
2 | 20 | 0.8±0.10b | 483±12b |
3 | 50 | 1.1±0.15a | 542±13a |
4 | 80 | 0.9±0.06ab | 490±15b |
5 | 100 | 0.8±0.15bc | 353±20c |
Treatments | AC (g·L-1) | Average seedling height (mm) | Average seedling biomass (mg) |
---|---|---|---|
1 | 0 | 7.6±0.15d | 1318±12d |
2 | 1 | 7.9±0.21d | 1434±18c |
3 | 1.5 | 9.8±0.06b | 1496±24b |
4 | 2 | 10.4±0.21a | 1612±23a |
5 | 2.5 | 9.9±0.15b | 1531±15b |
6 | 3 | 8.7±0.12c | 1423±18c |
Table 8 Effects of different concentrations of active charcoal (AC) on biomass amplification phase II
Treatments | AC (g·L-1) | Average seedling height (mm) | Average seedling biomass (mg) |
---|---|---|---|
1 | 0 | 7.6±0.15d | 1318±12d |
2 | 1 | 7.9±0.21d | 1434±18c |
3 | 1.5 | 9.8±0.06b | 1496±24b |
4 | 2 | 10.4±0.21a | 1612±23a |
5 | 2.5 | 9.9±0.15b | 1531±15b |
6 | 3 | 8.7±0.12c | 1423±18c |
Figure 2 HPLC image of taxane standards and Taxus tissue culture samples (A) HPLC of standard paclitaxel; (B) HPLC of sample paclitaxel; (C) HPLC of standard Baccatin III; (D) HPLC of sample Baccatin III; (E) HPLC of standard 10-DAB; (F) HPLC of sample 10-DAB
Paclitaxel (μg·g-1) | Baccatin III (μg·g-1) | 10-DAB (μg·g-1) | |
---|---|---|---|
Natural materials | 15.7 | 16.6 | 29.9 |
Tissue culture materials | 95.4 | 135.9 | 2036.0 |
Table 9 Contents of paclitaxel and taxane compounds
Paclitaxel (μg·g-1) | Baccatin III (μg·g-1) | 10-DAB (μg·g-1) | |
---|---|---|---|
Natural materials | 15.7 | 16.6 | 29.9 |
Tissue culture materials | 95.4 | 135.9 | 2036.0 |
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