In Vitro Propagation and Mutation Induction of Dendranthema Grandiflora Tzvelev
The genus Dendranthema (Family: Asteraceae) is a popular cut flower or pot plant species of high economic value cultivated around the world. The genus has more than 100 species of annuals and perennial herbs and shrubs used as floral crops as well as tea and source of other products such as pyret...
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| Format: | Thesis |
| Language: | English English |
| Published: |
2006
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| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/225/1/549054_FP_2006_12.pdf |
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| Summary: | The genus Dendranthema (Family: Asteraceae) is a popular cut flower or pot
plant species of high economic value cultivated around the world. The genus
has more than 100 species of annuals and perennial herbs and shrubs used as
floral crops as well as tea and source of other products such as pyrethrum. In
vitro propagation using meristems, shoot tips etc. have been successfully applied
as a means of large-scale production system for disease-free plants.
The present study is carried out in two parts: in vitro propagation and mutation
induction with the objectives of developing a protocol for an in vitro method of
propagation using ray florets and creation of variations in Dendranthema
grandiflora Tzvelev by combining the techniques of in vitro culture and radiationinduced
mutagenesis.In developing protocol for rapid propagation, in vitro culture was established by
using ray florets on two types of basal media, such as Murashige and Skoog
(MS) and Gamborg (B5) media, containing various levels of 6-benzylaminopurine
(BAP) (0, 0.5, 1.0 and 2.0 mg/L) and α–naphthaleneacetic acid (NAA) (0, 0.2,
0.5, 1.0 and 2.0 mg/L). In this study the highest percentage of callus formation
was observed after 8 weeks on MS medium supplemented with 2.0 mg/L BAP
and 1.0 mg/L NAA. However, the highest number of shoot multiplication was
obtained from MS basal medium containing 2.0 mg/L BAP and 1.0 mg/L NAA.
No growth responses were observed on MS and B5 basal media with BAP alone,
whereas some roots developed on NAA alone in both types of basal media.
After 10 weeks of culture many explants turned brown and died in the B5 basal
medium.
The highest callus proliferation in terms of fresh and dry weight as established on
MS basal media containing 2.0 mg/L BAP and 1.0 mg/L NAA followed by
treatment with 2.0 mg/L BAP with 1.0 and 2.0 mg/L NAA after 20 weeks of
culture. The highest number of adventitious shoot formation (6.0 shoots per
explant) was observed in MS medium supplemented with 2.0 mg/L BAP and 1.0
mg/L NAA. There were significant interactions between both growth regulators
on the number of shoot per explant and height of shoots produced.
In the radiation-induced mutagenesis study, irradiation treatments was performed
on callus derived from ray florets, and on fresh ray florets using gamma rays from 60Co source at levels of 0 Gy, 10 Gy, 20 Gy, 30 Gy, 40 Gy and 50 Gy at a dose
rate of 1.858 Gy/sec. Radiosensitivity test recorded a 100 % survival rate of
callus on control treatment and subsequent decrease on treatments at higher
doses. The highest growth of callus was observed in control treatment and
subsequently growth rate decreased correspond to increasing doses. Similarly,
irradiated ray florets recorded a 100% survival rate in the control treatment.
Treatments at 10, 20, 30, 40 and 50 Gy gave survival rates of 83.3%, 73.3%,
26.6%, 23.3% and 6.6% respectively.
The study concluded that the optimum dose for ray floret on growth rate of callus
and survival were between 18.8 - 28.4 Gy. The optimum dose for irradiated
callus based on growth rate and survival were between 26.9 - 36.2 Gy. The
results from the experiments indicated that mutation induction can be performed
on both ray florets and callus at LD50 18.8 - 28.4 Gy and 26.9 - 36.2 Gy
respectively. The formation of shoots was observed on control (0 Gy) and 10 Gy
treatments. The mean number of adventitious shoots from non-irradiated
samples (control) was 2.72 ± 0.09 compared to 2.5 ± 0.18 for treatment at 10 Gy.
No growth responses were observed from ray florets culture at all levels of
treatment. Therefore, the development of new variety through mutation induction
of Dendranthema, it is recommended to irradiate the explants at 10 Gy or lower.Amplified Fragment Length Polymorphism (AFLP) technique was carried out to
detect the variation on genomic DNA of callus samples irradiated at different
doses. The highest numbers of polymorphic bands were observed with primer
combinations E-AGC + M-CAG at 20 Gy and E-AGG + M-CAT at 10 Gy. The
smallest number of polymorphic bands was found with primer combination EAAG
+ M-CTC at 30 and 40 Gy. Primer combination E-ACG + M-CAA produced
the most number of polymorphic bands (80) when compared to other primer
combinations which were used in the experiment. There were no definite
correlation between the different levels of irradiation to the number of
polymorphic bands and unique polymorphic bands detected. Higher numbers of
polymorphic bands and unique polymorphic bands were detected by the various
primer combinations at doses 10 and 20 Gy. |
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