Chinese Bulletin of Botany ›› 2020, Vol. 55 ›› Issue (3): 318-328.DOI: 10.11983/CBB19211
• TECHNIQUE AND METHOD • Previous Articles Next Articles
Hong Luo,Xiaohui Wen,Yuanyuan Zhou,Silan Dai()
Received:
2019-10-28
Accepted:
2020-02-26
Online:
2020-05-01
Published:
2020-07-06
Contact:
Silan Dai
Hong Luo, Xiaohui Wen, Yuanyuan Zhou, Silan Dai. Establishment of In Vitro Regeneration System of Helenium aromaticum[J]. Chinese Bulletin of Botany, 2020, 55(3): 318-328.
Treatments | Plant growth regulators (mg·L-1) | |||
---|---|---|---|---|
NAA | 6-BA | 2,4-D | TDZ | |
A1 | 0.5 | 1 | 0 | 0 |
A2 | 0.5 | 2 | 1 | 0 |
A3 | 0.5 | 3 | 2 | 0 |
A4 | 1 | 1 | 1 | 0 |
A5 | 1 | 2 | 2 | 0 |
A6 | 1 | 3 | 0 | 0 |
A7 | 2 | 1 | 2 | 0 |
A8 | 2 | 2 | 0 | 0 |
A9 | 2 | 3 | 1 | 0 |
B1 | 0.2 | 1 | 0 | 0 |
B2 | 1 | 1 | 0 | 0 |
B3 | 0.2 | 2 | 0 | 0 |
B4 | 0.5 | 2 | 0 | 0 |
B5 | 1 | 2 | 0 | 0 |
B6 | 0.2 | 3 | 0 | 0 |
B7 | 0.5 | 3 | 0 | 0 |
C1 | 0.1 | 0.5 | 0 | 0.2 |
C2 | 0.2 | 1 | 0 | 0.2 |
C3 | 0.5 | 1.5 | 0 | 0.2 |
C4 | 0.2 | 0.5 | 0 | 0.5 |
C5 | 0.5 | 1 | 0 | 0.5 |
C6 | 0.1 | 1.5 | 0 | 0.5 |
C7 | 0.5 | 0.5 | 0 | 1 |
C8 | 0.1 | 1 | 0 | 1 |
C9 | 0.2 | 1.5 | 0 | 1 |
Table 1 Medium for callus induction and adventitious buds differentiation of Helenium aromaticum
Treatments | Plant growth regulators (mg·L-1) | |||
---|---|---|---|---|
NAA | 6-BA | 2,4-D | TDZ | |
A1 | 0.5 | 1 | 0 | 0 |
A2 | 0.5 | 2 | 1 | 0 |
A3 | 0.5 | 3 | 2 | 0 |
A4 | 1 | 1 | 1 | 0 |
A5 | 1 | 2 | 2 | 0 |
A6 | 1 | 3 | 0 | 0 |
A7 | 2 | 1 | 2 | 0 |
A8 | 2 | 2 | 0 | 0 |
A9 | 2 | 3 | 1 | 0 |
B1 | 0.2 | 1 | 0 | 0 |
B2 | 1 | 1 | 0 | 0 |
B3 | 0.2 | 2 | 0 | 0 |
B4 | 0.5 | 2 | 0 | 0 |
B5 | 1 | 2 | 0 | 0 |
B6 | 0.2 | 3 | 0 | 0 |
B7 | 0.5 | 3 | 0 | 0 |
C1 | 0.1 | 0.5 | 0 | 0.2 |
C2 | 0.2 | 1 | 0 | 0.2 |
C3 | 0.5 | 1.5 | 0 | 0.2 |
C4 | 0.2 | 0.5 | 0 | 0.5 |
C5 | 0.5 | 1 | 0 | 0.5 |
C6 | 0.1 | 1.5 | 0 | 0.5 |
C7 | 0.5 | 0.5 | 0 | 1 |
C8 | 0.1 | 1 | 0 | 1 |
C9 | 0.2 | 1.5 | 0 | 1 |
Treatments | Plant growth regulators (mg·L-1) |
---|---|
NAA | |
R0 | 0 |
R1 | 0.1 |
R2 | 0.2 |
R3 | 0.3 |
Table 2 Medium for root culture of Helenium aromaticum
Treatments | Plant growth regulators (mg·L-1) |
---|---|
NAA | |
R0 | 0 |
R1 | 0.1 |
R2 | 0.2 |
R3 | 0.3 |
Figure 1 The whole plant and flower morphology of Helenium aromaticum (A) The whole plant (Bar=2 cm); (B) The capitulum (Bar=2 mm); (C) Single flower (Bar=1 mm). PE: Petal; OV: Ovary; PI: Pistil; GT: Glandular trichomes
Figure 2 Several stages of callus induction and adventitious bud differentiation of Helenium aromaticum leaves (C2) (A) Leaf explants; (B) Calli (20 d, shown by arrow); (C) Adventitious buds (30 d, shown by arrow). Bars=1 cm. C2 is the same as Table 1.
Figure 3 Several stages of callus induction and adventitious bud differentiation of Helenium aromaticum stem fragments (B1) (A) Stem segment explants; (B) Calli (17 d, shown by arrow); (C) Adventitious buds (65 d, shown by arrow). Bars=1 cm. B1 is the same as Table 1.
Figure 4 Several stages of callus induction of Helenium aromaticum hypocotyls (C4) (A) Hypocotyl explants; (B) Calli (20 d). Bars=1 cm. C4 is the same as Table 1.
Explant types | Callus induction medium | Adventitious bud differentiation medium | Regeneration rate (%) | Rooting medium |
---|---|---|---|---|
Leaf | MS+0.2 mg·L-1 NAA+1 mg·L-1 6-BA+0.2 mg·L-1 TDZ | 62.1 | 1/2MS | |
Stem segment | MS+0.2 mg·L-1 NAA+1 mg·L-1 6-BA | 9.75 | 1/2MS | |
Hypocotyl | MS+0.2 mg·L-1 NAA+0.5 mg·L-1 6-BA+0.5 mg·L-1 TDZ | - | - | - |
Table 3 Different explant regeneration systems of Helenium aromaticum
Explant types | Callus induction medium | Adventitious bud differentiation medium | Regeneration rate (%) | Rooting medium |
---|---|---|---|---|
Leaf | MS+0.2 mg·L-1 NAA+1 mg·L-1 6-BA+0.2 mg·L-1 TDZ | 62.1 | 1/2MS | |
Stem segment | MS+0.2 mg·L-1 NAA+1 mg·L-1 6-BA | 9.75 | 1/2MS | |
Hypocotyl | MS+0.2 mg·L-1 NAA+0.5 mg·L-1 6-BA+0.5 mg·L-1 TDZ | - | - | - |
Treatments | Time of rooting (d) | The situation of root and plant | Rate of rooting (%) | Budding time (d) | Rate of flowering (%) |
---|---|---|---|---|---|
R0 | 16 | Root system is sparse and slender; the plant is robust with normal stems and leaves, flowering normally | 63.33 | 30 | 93.33 |
R1 | 10 | The root system is dense and crude, some of which are deformed roots; the plants are dwarf with large leaves | 45.46 | 34 | 86.36 |
R2 | 13 | The root system is few and short, most of which are deformed roots; the plants dwarf can’t grow with normal leaves | 12.50 | - | 0 |
R3 | 18 | Root deformity and plants hardly grow taller and leaves weaker | 14.71 | - | 0 |
Table 4 Rooting results of adventitious buds of Helenium aromaticum
Treatments | Time of rooting (d) | The situation of root and plant | Rate of rooting (%) | Budding time (d) | Rate of flowering (%) |
---|---|---|---|---|---|
R0 | 16 | Root system is sparse and slender; the plant is robust with normal stems and leaves, flowering normally | 63.33 | 30 | 93.33 |
R1 | 10 | The root system is dense and crude, some of which are deformed roots; the plants are dwarf with large leaves | 45.46 | 34 | 86.36 |
R2 | 13 | The root system is few and short, most of which are deformed roots; the plants dwarf can’t grow with normal leaves | 12.50 | - | 0 |
R3 | 18 | Root deformity and plants hardly grow taller and leaves weaker | 14.71 | - | 0 |
Figure 5 Rooting and plant status of Helenium aromaticum adventitious buds (from left to right are R0, R1, R2, and R3, respectively) (A1) Rooting condition (17 d); (A2) Plant status (17 d); (B1) Rooting condition (32 d); (B2) Plant status (32 d). Bars=1 cm. R0-R3 are the same as Table 2.
Figure 6 Establishment of in vitro regeneration system of Helenium aromaticum leaves (A) The leaf; (B) The leaf swelling (15 d); (C) Calli were induced from leaves (20 d); (D) Adventitious buds could be regenerated from calli (50 d); (E) Inducing rooting (60 d); (F) The adventitious roots induced after 16 days (76 d). Bars=2 cm
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