PROPAGATION OF CALLA (ZANTEDESCHIA SP.) PLANTS BY TISSUE CULTURE TECHNIQUE
NERMEEN EL–SAYED ABDUL–MONEEM ABDUL-QADER;
Abstract
This work was carried out in the Tissue Culture Laboratory, Horticulture Research Institute, Agricultural Research Center, Giza, Egypt during the period 2008 to 2013.
In-vitro produced explants of Zantedeschia ‘Hot Cherry” were obtained from the Tissue Culture Laboratory of Zohriya Garden, where they were subcultured on MS medium supplemented with BA at 2 ppm. However, explants remained unresponsive, lost their green color and became pale. To solve this problem, the effect of N, Fe and Mg levels in MS Medium were investigated. Further experiments on multiplication, rooting and tuberization were carried out. Results of all these experiments could be briefed in the following.
V.1. Exp: 1 - Effect of NH4NO3 level in MS medium
Raising the level of NH4NO3 in MS medium to 175% of the original one resulted in the highest values of total number of shoots and total number of leaves. Although raising the level in question to 200% gave rise to the highest shoot length, total fresh weight of shoots and chlorophyll content, the highest shoot length achieved could not be considered a privilege, as these shoots were abnormal.
V.2. Exp: 2 - Effect of FeSO4.7H2O level in MS medium on growth:
Using MS medium at 100% level of FeSO4.7H2O, i.e. the control treatment, gave rise to the highest records of number of shoots, total fresh weight of shoots and total number of leaves.
V.3. Exp: 3 - Effect of MgSO4.7H2O level in MS medium on growth:
Using 100, 150 or 175% of MgSO4.7H2O levels in MS medium gave rise to the heaviest propagules fresh weight. However, raising Mg levels above the normal one caused the multitude of shoots that were obtained at the normal level to turn to clumps of green callus.
V.4. Exp: 4 - Effect of cytokinin type, concentration and their interaction on multiplication
V.4.1. Effect of cytokinin type:
BA proved its superiority over Kin in achieving higher values of number of shoots, total fresh weight of shoots and total number of leaves. On the contrary, longer shoots were obtained when Kin was used.
V.4.2. Effect of cytokinin concentration:
Using cytokinins at 4 ppm resulted in the highest values of number of shoots, total fresh weight of shoots and total number of leaves, while the longest shoots were obtained at 2 ppm.
No shoots at all were produced on cytokinin free-media (concentration 0 ppm) in addition to media supplemented with cytokinins at 1 or 6 ppm, and explants died eventually.
V.4.3. Effect of the interaction between cytokinin type and cytokinin concentration:
Using BA at 4 ppm achieved the highest position for the characters of number of shoots, total fresh weight of shoots and total number of leaves. The treatment of Kin at 2 ppm also reached the first position for the character of shoot length.
It should be mentioned that media supplemented with either one of the two cytokinins at 0, 1, or 6 ppm, in addition to media enriched with BA at 5 ppm, failed to produce any number of shoots at all. Explants inoculated in these media died eventually.
V.5. Exp: 5 – Effect of auxin type, concentration and their interaction on rooting
V.5.1. Effect of auxin type: IBA surpassed NAA in regard to characters such as number of roots, root length, fresh weight of roots and percentage of rooting, though the difference between the two auxins was insignificant in case of the last character. On the other hand, NAA was superior when it concerns root diameter and number of days to rooting, though the difference between the two auxins was insignificant in case of the last character. However, this superiority should not be considered as a privilege, as thinner roots are functionally better and less days to rooting is a benefit.
V.5.2. Effect of auxin concentration:
Using auxins at 5 ppm scored the highest values of number of roots, root length, fresh weight of roots, root diameter and percentage of rooting. Using auxins at 6 ppm shared in the highest rank in case of root length, root diameter and percentage of rooting. Concentrations of 2, 3 and 4 ppm reached this first category only for the character of percentage of rooting, while concentration 0 ppm got a foothold in this position for number of days to rooting.
V.5.3. Effect of the interaction between auxin type and auxin concentration:
Using IBA at 5 ppm guaranteed the highest values in regard to number of roots, root length, fresh weight of roots and percentage of rooting. The least number of days to rooting was achieved by IBA at 5 ppm. However, this should be considered as a privilege to using IBA at 5 ppm.
V.6. Exp: 6 - Effect of incubation temperature and sucrose concentration and their interaction:
V.6.1. Effect of incubation temperature:
Mini-tubers produced by shoos incubated at 20°C outnumbered those produced on shoos incubated at 25°C. On the other hand, mini-tubers induced at 25°C surpassed those induced at 20°C concerning fresh weight and diameter.
V.6.2. Effect of sucrose concentration:
Enriching MS media with sucrose at 60 g/l resulted in the highest number of mini-tubers and fresh weight of mini-tubers. The highest position was also shared by treatment of sucrose at 70 g/l for characters of number of mini-tubers and mini-tuber diameter; and sucrose at 80 g/l for number of mini-tubers. No mini-tubers were produced at all when sucrose at 30 g/l was used.
V.6.3. Effect of the interaction between incubation temperature and sucrose concentration:
Adjusting incubation temperature to 20 °C resulted in the highest number of mini-tubers when sucrose level was 60, 70 or 80 g/l. On the other hand, raising incubation temperature to 25 °C gave rise to the highest fresh weight of mini-tubers at 60 g/l sucrose and to the highest mini-tuber diameter at 70 g/l sucrose.
The combination of sucrose at 30 g/l plus incubation temperature of either 20 or 25 °C yielded no mini-tubers at all.
V.7. Exp: 7 - Effect of sucrose and BA concentrations and their interaction:
V.7.1. Effect of sucrose concentration:
Using sucrose at 30 g/l gave rise to the highest records of number of shoots, shoot length, total fresh weight of shoots and total number of leaves. Using the same material at 60 g/l resulted in the highest shoot length, total fresh weight of shoots and percentage of tuber formation, while using it at 90 g/l achieved the highest value of number of mini-tubers and percentage of tuber formation.
V.7.2. Effect of BA concentration:
The highest values of number of shoots, total fresh weight of shoots, total number of leaves and number of mini-tubers were obtained when BA at 4 ppm was applied. Using BA at 8 ppm achieved also the highest position concerning shoot length, total fresh weight of shoots, number of mini-tubers, total fresh weight of mini-tubers, mini-tuber diameter and percentage of tuber formation.
Treatments deprived of BA failed to produce any shoots, and consequently no mini-tubers were obtained.
V.7.3. Effect of the interaction between sucrose and BA concentrations:
In-vitro produced explants of Zantedeschia ‘Hot Cherry” were obtained from the Tissue Culture Laboratory of Zohriya Garden, where they were subcultured on MS medium supplemented with BA at 2 ppm. However, explants remained unresponsive, lost their green color and became pale. To solve this problem, the effect of N, Fe and Mg levels in MS Medium were investigated. Further experiments on multiplication, rooting and tuberization were carried out. Results of all these experiments could be briefed in the following.
V.1. Exp: 1 - Effect of NH4NO3 level in MS medium
Raising the level of NH4NO3 in MS medium to 175% of the original one resulted in the highest values of total number of shoots and total number of leaves. Although raising the level in question to 200% gave rise to the highest shoot length, total fresh weight of shoots and chlorophyll content, the highest shoot length achieved could not be considered a privilege, as these shoots were abnormal.
V.2. Exp: 2 - Effect of FeSO4.7H2O level in MS medium on growth:
Using MS medium at 100% level of FeSO4.7H2O, i.e. the control treatment, gave rise to the highest records of number of shoots, total fresh weight of shoots and total number of leaves.
V.3. Exp: 3 - Effect of MgSO4.7H2O level in MS medium on growth:
Using 100, 150 or 175% of MgSO4.7H2O levels in MS medium gave rise to the heaviest propagules fresh weight. However, raising Mg levels above the normal one caused the multitude of shoots that were obtained at the normal level to turn to clumps of green callus.
V.4. Exp: 4 - Effect of cytokinin type, concentration and their interaction on multiplication
V.4.1. Effect of cytokinin type:
BA proved its superiority over Kin in achieving higher values of number of shoots, total fresh weight of shoots and total number of leaves. On the contrary, longer shoots were obtained when Kin was used.
V.4.2. Effect of cytokinin concentration:
Using cytokinins at 4 ppm resulted in the highest values of number of shoots, total fresh weight of shoots and total number of leaves, while the longest shoots were obtained at 2 ppm.
No shoots at all were produced on cytokinin free-media (concentration 0 ppm) in addition to media supplemented with cytokinins at 1 or 6 ppm, and explants died eventually.
V.4.3. Effect of the interaction between cytokinin type and cytokinin concentration:
Using BA at 4 ppm achieved the highest position for the characters of number of shoots, total fresh weight of shoots and total number of leaves. The treatment of Kin at 2 ppm also reached the first position for the character of shoot length.
It should be mentioned that media supplemented with either one of the two cytokinins at 0, 1, or 6 ppm, in addition to media enriched with BA at 5 ppm, failed to produce any number of shoots at all. Explants inoculated in these media died eventually.
V.5. Exp: 5 – Effect of auxin type, concentration and their interaction on rooting
V.5.1. Effect of auxin type: IBA surpassed NAA in regard to characters such as number of roots, root length, fresh weight of roots and percentage of rooting, though the difference between the two auxins was insignificant in case of the last character. On the other hand, NAA was superior when it concerns root diameter and number of days to rooting, though the difference between the two auxins was insignificant in case of the last character. However, this superiority should not be considered as a privilege, as thinner roots are functionally better and less days to rooting is a benefit.
V.5.2. Effect of auxin concentration:
Using auxins at 5 ppm scored the highest values of number of roots, root length, fresh weight of roots, root diameter and percentage of rooting. Using auxins at 6 ppm shared in the highest rank in case of root length, root diameter and percentage of rooting. Concentrations of 2, 3 and 4 ppm reached this first category only for the character of percentage of rooting, while concentration 0 ppm got a foothold in this position for number of days to rooting.
V.5.3. Effect of the interaction between auxin type and auxin concentration:
Using IBA at 5 ppm guaranteed the highest values in regard to number of roots, root length, fresh weight of roots and percentage of rooting. The least number of days to rooting was achieved by IBA at 5 ppm. However, this should be considered as a privilege to using IBA at 5 ppm.
V.6. Exp: 6 - Effect of incubation temperature and sucrose concentration and their interaction:
V.6.1. Effect of incubation temperature:
Mini-tubers produced by shoos incubated at 20°C outnumbered those produced on shoos incubated at 25°C. On the other hand, mini-tubers induced at 25°C surpassed those induced at 20°C concerning fresh weight and diameter.
V.6.2. Effect of sucrose concentration:
Enriching MS media with sucrose at 60 g/l resulted in the highest number of mini-tubers and fresh weight of mini-tubers. The highest position was also shared by treatment of sucrose at 70 g/l for characters of number of mini-tubers and mini-tuber diameter; and sucrose at 80 g/l for number of mini-tubers. No mini-tubers were produced at all when sucrose at 30 g/l was used.
V.6.3. Effect of the interaction between incubation temperature and sucrose concentration:
Adjusting incubation temperature to 20 °C resulted in the highest number of mini-tubers when sucrose level was 60, 70 or 80 g/l. On the other hand, raising incubation temperature to 25 °C gave rise to the highest fresh weight of mini-tubers at 60 g/l sucrose and to the highest mini-tuber diameter at 70 g/l sucrose.
The combination of sucrose at 30 g/l plus incubation temperature of either 20 or 25 °C yielded no mini-tubers at all.
V.7. Exp: 7 - Effect of sucrose and BA concentrations and their interaction:
V.7.1. Effect of sucrose concentration:
Using sucrose at 30 g/l gave rise to the highest records of number of shoots, shoot length, total fresh weight of shoots and total number of leaves. Using the same material at 60 g/l resulted in the highest shoot length, total fresh weight of shoots and percentage of tuber formation, while using it at 90 g/l achieved the highest value of number of mini-tubers and percentage of tuber formation.
V.7.2. Effect of BA concentration:
The highest values of number of shoots, total fresh weight of shoots, total number of leaves and number of mini-tubers were obtained when BA at 4 ppm was applied. Using BA at 8 ppm achieved also the highest position concerning shoot length, total fresh weight of shoots, number of mini-tubers, total fresh weight of mini-tubers, mini-tuber diameter and percentage of tuber formation.
Treatments deprived of BA failed to produce any shoots, and consequently no mini-tubers were obtained.
V.7.3. Effect of the interaction between sucrose and BA concentrations:
Other data
| Title | PROPAGATION OF CALLA (ZANTEDESCHIA SP.) PLANTS BY TISSUE CULTURE TECHNIQUE | Other Titles | إكثار نباتات الكلا بتقنية زراعة الأنسجة | Authors | NERMEEN EL–SAYED ABDUL–MONEEM ABDUL-QADER | Issue Date | 2014 |
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