In the present study, two accessions of Centella asiatica (CA03 and CA23) were subjected to gamma radiation to examine the response of these accessions in terms of survival rate, flavonoid contents, leaf gas exchange and leaf mass. Radiation Sensitivity Tests revealed that based on the survival rate, the LD50 (gamma doses that killed 50% of the plantlets) of the plantlets were achieved at 60 Gy for CA03 and 40 Gy for CA23. The nodal segments were irradiated with gamma rays at does of 30 and 40 Gy for Centella asiatica accession ‘CA03’ and 20 and 30 Gy for accession ‘CA23. The nodal segment response to the radiation was evaluated by recording the flavonoid content, leaf gas exchange and leaf biomass. The experiment was designed as RCBD with five replications. Results demonstrated that the irradiated plantlets exhibited greater total flavonoid contents (in eight weeks) significantly than the control where the control also exhibited the highest total flavonoid contents in the sixth week of growth; 2.64 ± 0.02 mg/g DW in CA03 and 8.94 ± 0.04 mg/g DW in CA23. The total flavonoid content was found to be highest after eight weeks of growth, and this, accordingly, stands as the best time for leaf harvest. Biochemical differentiation based on total flavonoid content revealed that irradiated plantlets in CA23 at 20 and 30 Gy after eight weeks contained the highest total flavonoid concentrations (16.827 ± 0.02; 16.837 ± 0.008 mg/g DW, respectively) whereas in CA03 exposed to 30 and 40 Gy was found to have the lowest total flavonid content (5.83 ± 0.11; 5.75 ± 0.03 mg/g DW). Based on the results gathered in this study, significant differences were found between irradiated accessions and control ones in relation to the leaf gas. The highest PN and gs were detected in CA23 as control followed by CA23 irradiated to 20Gy (CA23G20) and CA23G30 and the lowest PN and gs were observed in CA03 irradiated to 40Gy (CA03G40). Moreover, there were no significant differences in terms of PN and gs among the irradiated plants in each accession. The WUE of both irradiated accessions of Centella asiatica were reduced as compared with the control plants (p < 0.01) while Ci and E were enhanced. There were no significant differences in the gas exchange parameters among radiated plants in each accession. Moreover, malondialdehyde (MDA) of accessions after gamma treatments were significantly higher than the control, however, flavonoids which were higher concentration in irradiated plants can scavenge surplus free radicals. Therefore, the findings of this study have proven an efficient method of in vitro mutagenesis through gamma radiation based on the pharmaceutical demand to create economically superior mutants of C. asiatica. In other words, the results of this study suggest that gamma irradiation on C. asiatica can produce mutants of agricultural and economical importance.
Background: Chlorophytum borivilianum is a rare medicinal plant originally distributed throughout the forest of India. The tubers of C. borivilianum are used as an aphrodisiac and impotence supplement. The propagation of C. borivilianum is possible through seeds and tubers, but conventional methods may take several months. Hence in vitro technique of shoot regeneration could be an efficient alternative means of propagating the species. Latest study reported microtuberization of C. borivilianum but there is no sufficient study on a rapid method for shoot multiplication and elongation. Results: Young shoot buds of C. borivilianum were cultured on MS medium containing 6-benzylaminopurine (BAP) and Kinetin (Kn), both at 0, 8.88, 17.8 and 26.6 μM, either individually or in combinations. Proliferated shoots were subcultured on fresh medium of the same constituents on week 3 of culture for further shoot multiplication and elongation. BAP alone (8.88-26.6 μM) was significantly effective on shoot multiplication, while Kn alone (8.88-26.6 μM) was significantly effective on shoot elongation compared to the control containing MS basal medium without any plant growth regulator. However, combination of both cytokinins stimulated an interaction producing higher shoot number and shoot length compared to their individual application. Conclusions: The most suitable combination was 8.88 μM BAP + 8.88 μM Kn, reaching a mean shoot number of 10.83 and shoot length of 6.85 cm.
A number of parameters which have been reported to influence genetic transformation via Agrobacterium-mediated transformation method were evaluated to increase the frequency of transformation of cabbage (Brassica oleracea subsp. capitata) cv. KY Cross with AtHSP101 gene. The binary vector pCAMHSP was designed and mobilized into two Agrobacterium tumefaciens strains C58 and GV2260. The study was carried out on hypocotyl and shoot tip explants of cabbage cv. KY Cross. Transformation parameters optimized were pre-culture medium, acetosyringone application, bacterial density and inoculation time. The polymerase chain reaction (PCR) assay and production of mRNA of AtHSP101 gene were confirmed by reverse transcription-polymerase chain reaction (RT-PCR). The expression of LacZ gene in the transgenic plants also showed that it could be applied as a plant transformation reporter gene in genetic transformation studies. Multiple shoot regeneration of hypocotyl and shoot tip explants of cabbage after co-cultivation with Agrobacterium was optimized and medium containing 2 mg/L BAP was observed to be the best for shoot regeneration after cocultivation. In this study, 45% and 32.5% transformation efficiencies were achieved for hypocotyl and shoot tip explants, respectively using the optimized procedure.
The use of in vitro culture has been accepted as an efficient technique for clonal propagation of many woody plants. In the present research, we report the results of a number of experiments aimed at optimizing micropropagation protocol for tea (Camellia sinensis (L.) O. Kuntze) (clone Iran 100) using nodal segments as the explant. The effect of different combinations and concentrations of plant growth regulators (PGR) (BAP, TDZ, GA3) on shoot multiplication and elongation was assessed. The influence of exposure to IBA in liquid form prior to transfer to solid media on rooting of tea microshoots was investigated. The results of this study showed that the best treatment for nodal segment multiplication in terms of the number of shoot per explant and shoot elongation was obtained using 3 mg/L BAP in combination with 0.5 mg/L GA3. TDZ was found to be inappropriate for multiplication of tea clone Iran 100 as it resulted in hyperhydricity especially at concentrations higher than 0.05 mg/L. Healthy shoots treated with 300 mg/L IBA for 30 min followed by transfer to 1/2 strength MS medium devoid of PGR resulted in 72.3% of shoots producing roots and upon transferring them to acclimatization chamber 65% survival was obtained prior to field transfer.
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