Chinese Bulletin of Botany ›› 2023, Vol. 58 ›› Issue (3): 449-460.DOI: 10.11983/CBB22135
• TECHNIQUES AND METHODS • Previous Articles Next Articles
Minling Liao1, Ya Pu1, Xiaoyun Wu1, Chaofeng Ma1, Wenkui Wang2, Silan Dai1()
Received:
2022-06-28
Accepted:
2022-09-26
Online:
2023-05-01
Published:
2023-05-17
Contact:
*E-mail: silandai@sina.com
Minling Liao, Ya Pu, Xiaoyun Wu, Chaofeng Ma, Wenkui Wang, Silan Dai. Establishment of Regeneration System of Chrysanthemum indicum in Pingtan with Various Ligulate Floret Form[J]. Chinese Bulletin of Botany, 2023, 58(3): 449-460.
No. | Basal medium | 6-BA (mg?L-1) | NAA (mg?L-1) |
---|---|---|---|
Y1 | MS | 1.0 | 0.5 |
Y2 | MS | 1.0 | 1.0 |
Y3 | MS | 1.0 | 2.0 |
Y4 | MS | 2.0 | 0.5 |
Y5 | MS | 2.0 | 1.0 |
Y6 | MS | 2.0 | 2.0 |
Y7 | MS | 3.0 | 0.5 |
Y8 | MS | 3.0 | 1.0 |
Y9 | MS | 3.0 | 2.0 |
CK | MS | 0 | 0 |
Table 1 The composition of the medium for callus induction and adventitious buds differentiation
No. | Basal medium | 6-BA (mg?L-1) | NAA (mg?L-1) |
---|---|---|---|
Y1 | MS | 1.0 | 0.5 |
Y2 | MS | 1.0 | 1.0 |
Y3 | MS | 1.0 | 2.0 |
Y4 | MS | 2.0 | 0.5 |
Y5 | MS | 2.0 | 1.0 |
Y6 | MS | 2.0 | 2.0 |
Y7 | MS | 3.0 | 0.5 |
Y8 | MS | 3.0 | 1.0 |
Y9 | MS | 3.0 | 2.0 |
CK | MS | 0 | 0 |
No. | Basal medium | NAA (mg?L-1 ) | 6-BA (mg?L-1 ) | TDZ (mg?L-1 ) |
---|---|---|---|---|
T1 | MS | 2.0 | 1.0 | 0.1 |
T2 | MS | 2.0 | 1.0 | 0.2 |
T3 | MS | 2.0 | 1.0 | 0.4 |
T4 | MS | 2.0 | 2.0 | 0.1 |
T5 | MS | 2.0 | 2.0 | 0.2 |
T6 | MS | 2.0 | 2.0 | 0.4 |
T7 | MS | 2.0 | 3.0 | 0.1 |
T8 | MS | 2.0 | 3.0 | 0.2 |
T9 | MS | 2.0 | 3.0 | 0.4 |
Table 2 The composition of the medium for callus induction with different concentrations of TDZ
No. | Basal medium | NAA (mg?L-1 ) | 6-BA (mg?L-1 ) | TDZ (mg?L-1 ) |
---|---|---|---|---|
T1 | MS | 2.0 | 1.0 | 0.1 |
T2 | MS | 2.0 | 1.0 | 0.2 |
T3 | MS | 2.0 | 1.0 | 0.4 |
T4 | MS | 2.0 | 2.0 | 0.1 |
T5 | MS | 2.0 | 2.0 | 0.2 |
T6 | MS | 2.0 | 2.0 | 0.4 |
T7 | MS | 2.0 | 3.0 | 0.1 |
T8 | MS | 2.0 | 3.0 | 0.2 |
T9 | MS | 2.0 | 3.0 | 0.4 |
No. | Basal medium | NAA (mg?L-1 ) |
---|---|---|
G1 | 1/2MS | 0.2 |
G2 | 1/2MS | 0.5 |
G3 | 1/2MS | 1.0 |
1/2MS | 1/2MS | 0 |
MS | MS | 0 |
Table 3 Formula of rooting medium
No. | Basal medium | NAA (mg?L-1 ) |
---|---|---|
G1 | 1/2MS | 0.2 |
G2 | 1/2MS | 0.5 |
G3 | 1/2MS | 1.0 |
1/2MS | 1/2MS | 0 |
MS | MS | 0 |
Figure 1 Ray floret variations in Chrysanthemum indicum in Pingtan (A1) Flat type; (A2) Spoon type; (A3) Tubular type; (A4) Mix type; (B1), (B2) Chenille-like; (C1), (C2) Swallowtail-like; (D1) Evaginable; (D2) Incurvate; (E1) Difference in ray floret length; (E2), (E3) Difference in degree of curvature; (E4) Difference in cracking degree; (F1)-(F4) Difference in ray floret number; (G1)-(G4) Difference in proportion of each petal type in mix type; (H) Difference in diameter of capitulum. Bars=1 cm
Figure 2 Chrysanthemum indicum in Pingtan stains with various ligulate floret form (A) Plant (bar=3.5 cm); (B) Leaf shape (bar=1 cm); (C) Capitulum (bar=1 cm)
Treat-ments | Callus formation rate on the 14th day (%) | Differentiation rate on the 40th day (%) | Treatments | Callus formation rate on the 14th day (%) | ||
---|---|---|---|---|---|---|
Leaves | tTCLs | Leaves | tTCLs | |||
Y1 | 100.00±0.00 a | 100.00±0.00 a | 0 | 82.00±3.46 a | T1 | 100.00±0.00 a |
Y2 | 100.00±0.00 a | 100.00±0.00 a | 0 | 53.50±4.95 c | T2 | 100.00±0.00 a |
Y3 | 100.00±0.00 a | 90.50±4.95 b | 3.71±0.19 b | 33.50±6.36 d | T3 | 97.00±0.04 ab |
Y4 | 100.00±0.00 a | 100.00±0.00 a | 0 | 63.50±4.95 bc | T4 | 83.50±0.06 ab |
Y5 | 100.00±0.00 a | 100.00±0.00 a | 0 | 67.00±5.66 b | T5 | 97.50±0.03 ab |
Y6 | 100.00±0.00 a | 98.50±2.12 a | 9.39±3.35 b | 57.50±0.71 bc | T6 | 87.50±0.09 ab |
Y7 | 100.00±0.00 a | 100.00±0.00 a | 0 | 61.00±1.41 bc | T7 | 81.50±0.05 b |
Y8 | 100.00±0.00 a | 93.50±0.71 b | 0 | 16.50±3.54 e | T8 | 85.00±0.14 ab |
Y9 | 75.48±4.30 b | 98.50±2.12 a | 19.09±1.29 a | 41.00±4.24 d | T9 | 90.00±0.06 ab |
CK | 0.00±0.00 c | 0.00±0.00 c | 0 | 0 | - | - |
Table 4 Effects of different media on callus induction and adventitious buds differentiation for leaves and transverse thin cell layers (tTCLs) of Chrysanthemum indicum in Pingtan
Treat-ments | Callus formation rate on the 14th day (%) | Differentiation rate on the 40th day (%) | Treatments | Callus formation rate on the 14th day (%) | ||
---|---|---|---|---|---|---|
Leaves | tTCLs | Leaves | tTCLs | |||
Y1 | 100.00±0.00 a | 100.00±0.00 a | 0 | 82.00±3.46 a | T1 | 100.00±0.00 a |
Y2 | 100.00±0.00 a | 100.00±0.00 a | 0 | 53.50±4.95 c | T2 | 100.00±0.00 a |
Y3 | 100.00±0.00 a | 90.50±4.95 b | 3.71±0.19 b | 33.50±6.36 d | T3 | 97.00±0.04 ab |
Y4 | 100.00±0.00 a | 100.00±0.00 a | 0 | 63.50±4.95 bc | T4 | 83.50±0.06 ab |
Y5 | 100.00±0.00 a | 100.00±0.00 a | 0 | 67.00±5.66 b | T5 | 97.50±0.03 ab |
Y6 | 100.00±0.00 a | 98.50±2.12 a | 9.39±3.35 b | 57.50±0.71 bc | T6 | 87.50±0.09 ab |
Y7 | 100.00±0.00 a | 100.00±0.00 a | 0 | 61.00±1.41 bc | T7 | 81.50±0.05 b |
Y8 | 100.00±0.00 a | 93.50±0.71 b | 0 | 16.50±3.54 e | T8 | 85.00±0.14 ab |
Y9 | 75.48±4.30 b | 98.50±2.12 a | 19.09±1.29 a | 41.00±4.24 d | T9 | 90.00±0.06 ab |
CK | 0.00±0.00 c | 0.00±0.00 c | 0 | 0 | - | - |
Figure 3 Regeneration of Chrysanthemum indicum in Pingtan leaves in different media (A) Leaves in different media; (B) Leaves in media with TDZ. Y1-Y9, and CK are the same as in Table 1; T1-T9 are the same as in Table 2. Bars=4.5 cm
Figure 4 Adventitious bud differentiation for leaves of Chrysanthemum indicum in Pingtan in different media (A) Y3; (B) Y6; (C) Y9. Y3, Y6 and Y9 are the same as in Table 1. From left to right: Adventitious bud differentiation of leaves in different media at 21, 28, 35, 42 and 49 d. Bar=1 cm
Treatments | Browning rate on the 21th day (%) | Treat- ments | Browning rate on the 24th day (%) |
---|---|---|---|
Y1 | 83.04±7.92 abcd | T1 | 52.78±3.93 f |
Y2 | 96.43±5.05 a | T2 | 56.08±3.88 ef |
Y3 | 80.36±6.84 bcde | T3 | 81.80±0.78 c |
Y4 | 90.11±8.58 abc | T4 | 72.14±2.18 d |
Y5 | 77.56±3.95 cde | T5 | 89.45±0.78 b |
Y6 | 67.86±9.22 e | T6 | 61.51±0.56 e |
Y7 | 74.22±2.49 de | T7 | 97.50±3.54 a |
Y8 | 42.50±5.00 f | T8 | 100.00±0.00 a |
Y9 | 34.17±1.18 f | T9 | 87.30±2.25 bc |
CK | 92.73±9.50 ab |
Table 5 The browning rate for leaves of Chrysanthemum indicum in Pingtan in different media
Treatments | Browning rate on the 21th day (%) | Treat- ments | Browning rate on the 24th day (%) |
---|---|---|---|
Y1 | 83.04±7.92 abcd | T1 | 52.78±3.93 f |
Y2 | 96.43±5.05 a | T2 | 56.08±3.88 ef |
Y3 | 80.36±6.84 bcde | T3 | 81.80±0.78 c |
Y4 | 90.11±8.58 abc | T4 | 72.14±2.18 d |
Y5 | 77.56±3.95 cde | T5 | 89.45±0.78 b |
Y6 | 67.86±9.22 e | T6 | 61.51±0.56 e |
Y7 | 74.22±2.49 de | T7 | 97.50±3.54 a |
Y8 | 42.50±5.00 f | T8 | 100.00±0.00 a |
Y9 | 34.17±1.18 f | T9 | 87.30±2.25 bc |
CK | 92.73±9.50 ab |
Figure 5 Adventitious bud differentiation of transverse thin cell layers of Chrysanthemum indicum in Pingtan in different media (40 d) Y1-Y9 are the same as in Table 1. Bar=1 cm
Figure 6 Rooting and plant status of Chrysanthemum indicum in Pingtan adventitious buds (A1)-(E1) Plant status (0 d); (A2)-(E2) Rooting condition (10 d); (A3)-(E3) Plant status (26 d); (A4)-(E4) Rooting condition (26 d). From left to right: MS, 1/2MS, G1, G2, and G3 (the same as in Table 3). Bars=1 cm
Treat- ments | Time of rooting (d) | Rate of rooting (%) | Average root length (cm) | The situation of root and plant | Number of branches |
---|---|---|---|---|---|
MS | 6 | 66.7 | 12.59±2.60 a | The root system is dense, slender, and long; the plant is weak with branches and small leaves | 3±1 a |
1/2MS | 6 | 100 | 5.49±0.86 b | The root system is sparse and slender; the plant is weak with many branches and small leaves | 6±2 a |
G1 | 14 | 100 | 4.41±0.80 b | The root system is crude and short, with many root hairs; the plant is weak with branches | 4±2 a |
G2 | 10 | 100 | 3.21±2.62 b | The root system is crude and short, with many root hairs; the plant is robust and higher, with larger leaves | 2±1 a |
G3 | 24 | 100 | 3.80±2.22 b | The root system is crude and short, with many root hairs; the plant is robust and highest, with largest leaves | 2±1 a |
Table 6 Rooting and plant status of Chrysanthemum indicum in Pingtan adventitious buds
Treat- ments | Time of rooting (d) | Rate of rooting (%) | Average root length (cm) | The situation of root and plant | Number of branches |
---|---|---|---|---|---|
MS | 6 | 66.7 | 12.59±2.60 a | The root system is dense, slender, and long; the plant is weak with branches and small leaves | 3±1 a |
1/2MS | 6 | 100 | 5.49±0.86 b | The root system is sparse and slender; the plant is weak with many branches and small leaves | 6±2 a |
G1 | 14 | 100 | 4.41±0.80 b | The root system is crude and short, with many root hairs; the plant is weak with branches | 4±2 a |
G2 | 10 | 100 | 3.21±2.62 b | The root system is crude and short, with many root hairs; the plant is robust and higher, with larger leaves | 2±1 a |
G3 | 24 | 100 | 3.80±2.22 b | The root system is crude and short, with many root hairs; the plant is robust and highest, with largest leaves | 2±1 a |
Figure 7 The capitulum of regenerated plants and the proportion of each petal type in Chrysanthemum indicum in Pingtan (A1), (A2) Plant status after transplanting for 60 days, from left to right, their rooting media were MS, 1/2MS, G1, G2, and G3, respectively (bars=3.5 cm); (B) The capitulum of regenerated plants (bar=1 cm); (C) The proportion of flat petals, spoon petals and tubular petals in each capitulum; (D) Average proportion of each petal type in all capitulums. MS, 1/2MS, G1, G2, and G3 are the same as in Table 3.
Figure 8 Regeneration system of transverse thin cell layers in Chrysanthemum indicum in Pingtan stains with various ligulate floret forms (A), (B) Callus induction and adventitious bud differentiation in Y1; (C) Adventitious buds grown to 0.8 cm; (D), (E) Rooting and plant status of adventitious buds in G2; (F), (G) Plant status after transplanting for 30 and 90 days; (H) Flower buds; (I) Capitulum of regenerated plant. Y1 is the same as in Table 1; G2 is the same as in Table 3. Bars=2 cm
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