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BACKGROUND: In order to produce an effective callus in Echinacea purpurea L.; determination of the explant type and growth regulators that best respond to callus induction and the optimization of the culture conditions to increase the amount of caffeic acid derivatives (CADs) in the obtained callus. CADs contents of callus cultures of E. purpurea were evaluated by establishing an effective callus induction system in vitro. RESULTS: Various medium containing different growth regulators were tested using leaf, petiole, cotyledon and root as the explants. The best callus development was achieved in MS medium with 1.0 mg l 1 2,4- D + 2.0 mg l 1 BAP in leaf, 1.0 mg l 1 NAA + 0.5 mg l 1 TDZ in petiole, 2.0 mg l 1 NAA + 1.0 mg l 1 TDZ in cotyledon and 0.5 mg l 1 NAA + 0.5 mg l 1 BAP in roots. Upon optimisation of callus growth, each type of explant was cultured for 4, 6, 8 and 10 weeks in medium for the analyses of caftaric acid, chlorogenic acid, caffeic acid and chicoric acid contents. The highest amounts of caftaric acid (4.11 mg/g) and chicoric acid (57.89 mg/g) were found from petiole explants and chlorogenic acid (8.83 mg/g) from root explants at the end of the 10-week culture time. CONCLUSIONS: As a result of the present study, the production of caffeic acid derivatives was performed by providing the optimization of E. purpurea L. callus cultures. Effective and repeatable protocols established in this study may offer help for further studies investigating the production of caffeic acid derivatives in vitro.
Subject(s)
Caffeic Acids , Echinacea , Plant Growth Regulators , Time Factors , In Vitro Techniques , Cells, Cultured , Plant Roots/growth & development , Plant Leaves/growth & development , Cotyledon/growth & development , Culture TechniquesABSTRACT
Present study was conducted to standardize callus development and indirect organogenesis from differentexplants of Vernonia anthelmintica (L.) Willd. Among, the various hormonal combinations tested: IndoleAcetic Acid (IAA) and BA and IAA and Kinetin combinations were found to be optimum for inducing callusingwithin a time span of 10 days. Best callus response was observed in 1.5 mg l−1 IAA and 1.5 mg l−1 BA,which produced white friable callus. Best indirect shoot organogenesis was observed in 4 mg l−1 BA and6 mg l−1 kinetin. Elongated shoots when transferred on to half strength Murashige and Skoog (MS) Mediumsupplemented with Indole-3-butyric acid (IBA), rooting was observed. MS medium fortified with 2 mg l−1 IBAshowed better rooting than all other concentrations tested. This concentration produced maximum number ofroots and maximum percentage of rooting. Plantlets developed by indirect organogenesis of V. anthelminticawere successfully acclimatized to field conditions
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In the present study an attempt has been made to evaluate the phytochemical, antimicrobial, antioxidant and α-amylase inhibitory activity of Coccinia indica(W. and A) leaf extracts using four solvents and compare it with the callus extracts. Callus was initiated from the leaf explants of C.indicawith 90% efficiency using MS medium supplemented with BAP (1 mg/l) + NAA (0.2 mg/l). Successive extraction method of C.indicawas found to be an efficient method of extraction and methanol was observed to be the best suited solvent for the extraction of phytochemicals and macromolecules that were responsible for antimicrobial, antioxidant and α-amylase inhibition. GC-MS analysis of C.indicahas confirmed the presence of bioactive compounds (Example: 9-Octadecanoic acid, 2-octadecycloxy ethyl ester (100%) in successive methanolic callus extract) in all the extracts where the FTIR analysis has confirmed the presence of various important functional groups of the identified bioactive compounds. Successive methanol extract of callus of C.indicawas found to be the potent antimicrobial agent with drug efflux pump inhibitor property against 5 bacterial strains, Klebsiella pneumoniae (ATCC700603), Escherichia coli (ATCC25922), Staphylococcus aureus (ATCC 25923), Proteus mirabilis (ATCC 25933)and Pseudomonas aeruginosa (clinical isolate) and 3 fungal strains, Candida albicans (IFM 40009), Candida tropicalis (IFM 55058)andCandida krusei (IFM 46521).Successive methanol extract of callus of C.indicawas found to be an efficient antioxidant agent and an efficient α-amylase inhibitor, which proves it to be a potent anti-diabetic agent with IC50 concentration to be 82.5μg/ml. This study is one of the strong evidence for this plant to be used by the traditional practitioners as a phytopharmaceutical agent
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The study was carried out for callus induction and synthetic seed development from the shoot tips of Draceana sanderiana sander ex Mast. The shoot tips were subjected to different concentrations (0.25, 0.5 &1.0 mg/l) of 2,4-D on MS medium. The research findings revealed that the 2,4-D at concentrations of 0.25 mg/l was more suited for the profuse callus formation. The friable and light yellow callus was induced within 2 weeks of culture at 0.25 mg/l of 2,4-D on MS medium as compared to the other two concentrations of 2,4-D i.e.; 0.5 and 1.0 mg/l. Similarly the effect of sodium alginate and calcium chloride percentage on synthetic seed formation was observed, it was found that somatic embryos formed from shoot tips via callus kept in 2.5% sodium alginate and 100 milli molar CaCl2 produced synthetic seeds with firm spherical beads. The study leads to the formation of synthetic seeds of Draceana sanderiana which can be used for the conservation of germplasm through cryopreservation and the micro propagation of the said plant species.
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Young petiole of Tussilago farfara was used as the material to investigate the plant growth regulators which could influence in vitro culture and plant regeneration and to establish rapid propagation technique. The ideal sterilization method was that young petiole of T. farfara was sterilized with 75% ethanol for 30 s, and then transferred to saturated bleaching power supernatant for 15 min. The suitable medium for callus induction was MS+6-BA 3.0 mg•L⁻¹+2,4-D 2.0 mg•L⁻¹ with 96.2% induction rate. The seedlings had better differentiation with 91% differentiation rate and 8.26 buds on the medium containing MS+ZT 2.0 mg•L⁻¹+NAA 0.3 mg•L⁻¹. The preferred enrichment medium of adventitious bud was MS+KT 1.0 mg•L⁻¹+IBA 0.3 mg•L⁻¹ with 11.81 enrichment times and 4.9 cm seedling height. The rooting medium included 1/2MS+IBA 0.2 mg•L⁻¹ with the average number of rooting was 5.86 and the rooting rate was above 95.22%. The container seedlings can grow well and the survival rate was more than 90% when they were transplanted on the medium added with river sand and organic fertilizer with the ratio of 3∶1. The field experiments indicated that significant differences in increment and yield of pollen grains among the tissue-culture, cultivation and wild type of T. farfara under the same cultivation conditions. The cultivated plants were relatively high on the increment and yield of pollen grains. The active ingredient content of the tissue culture and the wild materials was basically the same.
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Objective: To induce callus from the medicinally valuable species, Barringtonia racemosa L. (B. racemosa) whereby the formation of callus is essential for micropropagation studies and in vitro plant secondary metabolites production. Methods: The callus induction potential in B. racemosa was assessed from endosperm explant cultured on different culture media and plant hormonal treatments. Lloyd and McCown's woody plant medium and Murashige and Skoog's medium were used in the study as culture media. On the other hand, various concentrations and combinations of 2,4-dichlorophenoxyacetic acid (1.0-2.0 mg/L) and kinetin (0.5-2.5 mg/L) had been incorporated in the culture media to exert the effects of auxin and cytokinin on callus induction. Results: From the present study, it was found that the profuse [(1.681 ± 0.770) g fresh weight, (0.239 ± 0.239) g dry weight] and friable callus formation was optimally produced with desirable morphology and considerable percentage of callus induction (56.70%) in endosperm explants cultured on 1.0 mg/L 2,4-dichlorophenoxyacetic acid and 1.5 mg/L kinetin in Murashige and Skoog's medium. Conclusions: A reliable protocol for inducing callus formation of profuse and friable morphology in endosperm explant of B. racemosa had therefore been successfully established.
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A protocol was developed for the micropropagation of Plumbago scandens L. from the shoot tip and node explants.The best response of shoot elongation (10.18±2.01 mm) was observed on MS basal medium supplemented with 0.02 mg/L IAA – 0.02 mg/L GA3. The maximum number of root induction (10.0±2.21) and shoot elongation (8.24±3.24 mm) was observed on medium containing 0.01 mg/L IBA and 0.01 mg/L GA3. The in vitro propagated plants were transferred to soil with 80% survival rate. Profuse compact callus was induced and proliferated from several explants (cotyledons, internodes, hypocotyls and roots) cultured on MS medium supplemented with all the combinations of 2,4-D – GA3 or 2,4-D alone and combinations of IAA – BAP or IAA alone, and the highest percentage of friable callus (90%) were induced in the sections of compact callus using 2.0 mg/L IAA – 0.02 mg/L BAP – 0.5 mg/L GA3.The qualitative determination of chemical constituents in the extracts was evaluated by a gas chromatography coupled to a mass spectrometry, and it was verified the presence of plumbagin only in root extracts but not in in vitro plantlets.The antibacterial activity of root extracts against various pathogenic bacteria, and the minimum inhibitory concentrations (MICs) was determined. Chloroform extracts showed good antibacterial activity against Neisseria gonorrhoeae between 0.4 to 1.0 mg/L with 20.4 to 30.0 mm (diameter zone of inhibition); inhibition against Staphylococcus aureus was moderate, and lower against Escherichia coli. Chloroform extracts had the lowest MICs for N. gonorrhoeae (<0.1 mg/mL per disc), and the activities against S. aureus (MIC 0.2 mg/mL) and E. coli (MIC 0.4 mg/mL) were less pronounced.
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Objective To study the conditions of callus induction with the roots of Aquilaria sinensis as explants. Methods Two sources of roots of Aquiliaria sinensis were selected as the explants. The effects of sterilization methods and the combination of different concentrations of phytohormones on callus induction were evaluated. Results When Aquiliaria sinensis root seedling was sterilized in 0.01mg/mL HgCl2 solution for 3 minutes, the sterilized effect was the best. The optimal callus induction medium was MS+0.1 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) +0.1 mg/L 6-benzylaminopurine (6-BA). Aseptic Aquiliaria sinensis root seedling cultivated in callus induction medium containing MS+1.0 mg/L naphthalene acetic acid ( NAA) +0.8 mg/L 6-BA achieved the highest callus induction rate. Conclusion Callus can be induced from two sources of Aquilaria sinensis roots. The induction rate of callus is lower when the explant root seedling is cultivated using 2,4-D alone as inducer, and is increased when used together with 6-BA.
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Objective: To optimize the proper condition for callus induction of Asarum heterotropoides var. mandshuricum. Methods: The effects of different explants, medium types, and plant growth regulators on the callus induction of A. heterotropoides var. mandshuricum were studied by using the petioles and leaves as explants. Also we aimed to discover the morphology characteristics of callus in different periods through paraffin method. Results: The efficient callus induction medium was 1/2 MS + 0.6 mg/L 6-BA + 0.15 mg/L NAA while using petioles and leaves during young period as explants. Callus induction from young leaves lasted so long time. Furthermore it was found that the induced calli were embryogenic for they were composed of compact, small and irregular arrangement cells. Subsequently these calli could be differentiated to develop the regeneration plantlet. Conclusion: Young petiole is the optimal explant for callus induction of A. heterotropoides var. mandshuricum, and the efficient callus induction condition we optimized would provide a basis for establishing the subsequent regeneration system.
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Objective: To study the combination and proportion for the explants in NaClO and the suitable hormone induced by callus of Fritillaria ebeiensis var. purpurea. The total alkaloids content variations of raw herb and callus were investigated. Methods: The fresh bulbs of F. ebeiensis var. purpurea were used as the exprimental materials and NaClO and antibiotics as disinfectants. It was cultured in vitro in MS medium with plant hormones 2,4-D + 6-BA and 2,4-D + KT by orthogonal test design. Alkaloids contents in original bulb and callus were measured with UV. Results: The best way to sterilization was at a concentration of 5% NaClO with 8 min. The optimal hormone levels was 2,4-D 0.5 mg/L + 6-BA 2.0 mg/L. The alkaloid contents of raw herbs and callus were 1.46% and 1.09%. Conclusion: Callus induced by tissue culture technique are soft, luster, and transparent, which are suitable for subculture. Total alkaloids content in callus is less than that in raw herb.
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Aims: The genetic improvement of garlic can be achieved by biotechnological manipulations as breeding in this vegetatively propagated crop is limited. The current research was conducted with a view to develop an efficient in vitro regeneration protocol for four local garlic accessions namely, G121, G122, G123 and G124. Place, Duration and Design of Study: The experiment was conducted in the Tissue Culture Laboratory of the Department of Genetics and Plant Breeding, Bangladesh Agricultural University during the period from June 2013 to June 2014 using three-factorial experimental design. Methodology: The root tips, basal disc and leaf base were cultured in MS medium supplemented with 2, 4-Dichlorophenoxyacetic acid (2, 4-D) alone, and with both 2, 4-D and 6-Benzylaminopurine (BAP) together for callus induction and the later for subsequent sub-culturing and proliferation of callus. MS medium supplemented with 1-Naphthaleneacetic acid (NAA) and BAP was used for plantlet regeneration. Results: The percentage of callus induction increased with the increase in the concentration of 2,4- D, starting from 0.5 mg L-1 till 2.0 mg L-1 and declined with further increase in the concentration of 2,4-D. The MS medium supplemented with 2,4-D and BAP showed higher percentage of callus induction and callus proliferation compared to that of with 2,4-D alone. The highest percentage of callus induction was observed in the genotype G124 from the explant basal disc (85%) and in the genotype G121 from the explant leaf base (80%) with 2.0 mg L-1 2,4-D and 2.0 mg L-1 BAP. MS medium supplemented with 2 mg L-1 2,4-D + 0.5 mg L-1 BAP showed highest percentage of callus proliferation (90%) in almost all the genotypes. The highest percentage of plantlet regeneration were observed in the genotype G124 for the explants basal disc (63.33%) in MS medium supplemented with 2 mg L-1 NAA + 1 mg L-1 BAP. The survival rate of the plantlets after acclimatization varied from 40% (in G123) to 70% (in G121). Conclusion: The optimized protocol of plant regeneration from local garlic accessions will be useful for any future garlic improvement programs using biotechnological means.
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Aims: A procedure was developed for embryogenesis from embryo explants derived from mature seeds of freshly harvested Serenut 4T, Serenut 1R and Acholi-white groundnut cultivars representing the three broad groundnut botanical classifications. Methodology: This study explored the use of mature embryo axes as explants for somatic embryogenesis, and determined the factors that affect regeneration of three Ugandan groundnut cultivars. Freshly harvested mature seeds of the three groundnut cultivars were collected and the embryo explants were initiated on 3 media namely; Murashige and Skoog (MS) basal media with varying concentrations of the growth regulator 2,4-Dichlorophenoxy acetic acid (2,4-D); Chu N6 basal medium with vitamins (N6); and Callus Induction Medium (CIM). The shoot formation and elongation medium contained MS basal medium supplemented with indolebutyric acid (IBA) and 6- Benzylamminopurine (BAP) in isolation, and BAP in combination with a-naphthaleneacetic acid (NAA) and indoleacetic acid (IAA). For root induction, elongated shoots were transferred to MS medium supplemented with various combinations of NAA with IBA, BAP and a combination of IBA and Kinetin. Results and Conclusion: Different concentrations of 2,4-D elicited different callogenesis responses in the cultivars with Acholi white (Valencia botanical) and Serenut 4T (Spanish botanical) giving the optimal response at 5mg/l whereas Serenut 1R (Virginia botanical) showed best response at a concentration of 30 mg/l. N6 and CIM supported callogenesis in Acholi white (AW) and Serenut 4T only. In all cultivars, maximum root production was gained when using MS medium supplemented with NAA- 1 mg/l and IBA -2.0 mg/l. On the other hand, for Serenut 1R and Serenut 4T, BAP 2.5 mg/l; NAA 0.5 mg/l combination yielded higher shoot regeneration percentage whereas for AW BAP 3 mg/l; NAA 0.5 mg/l supported maximum shoot production.This is the first ever report of successful regeneration of the three groundnuts botanicals in Uganda. These results are likely to facilitate genetic transformation of three preferred Ugandan groundnut varieties.
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In vitro stem segments of Pelargonium radula cultured for callusing then differentiated into somatic embryos and subsequently regenerated plantlets. Initiation of callus was observed in culture medium containing low concentrations of the plant growth regulators, 2,4-dichlorophenoxyacetic acid (2,4-D) and/or α-naphthalene acetic acid (NAA). At 0.2 mg/L 2,4-D and 0.2 mg/L NAA was showing the highest rate (92%) of callus induction. The callus showed no sign of browning after sub-cultured. Sub-culturing the callus onto medium with 0.2 mg/L 2,4-D showed the highest in proliferation rate resulted 13.45g weight of callus. The presence of agar at 6 g/L and 0.5 mg/L Gibberellic acid (GA3) improved the regeneration of the somatic embryos, which produced maximum number of plantlets (15 plantlets). Agar with concentration of 9 to12 g/L increased the incidence of browning. The former medium was more successful in plantlet regeneration when the selected embryos were subsequently transferred to regeneration medium with 3 g/L agar, 0.5 mg/L GA3 and 0.5 mg/L Benzylaminopurine (BAP).
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The seeds of C. nervosa and E. pseudoclavicaulis were germinated asymbiotically on Knudson C (KC) and Schenk and Hildebrandt basal medium (SH). Growth regulators such as 2,4-Dichlorophenoxyacetic acid (2,4-D) individually and in combinations with benzyladenine (BA) and kinetin were used for callus induction from the protocorm like bodies. Coelogyne nervosa showed maximum (90%) callus induction in Knudson C medium supplemented with 2,4-D (2.26 µM) and Eria pseudoclavicaulis showed 60% callus induction in Schenk and Hildebrandt medium supplemented with 2,4-D (2.26 µM). Calli developed a route of production of protocorm-like bodies and eventually developed into plantlets on transfer to growth regulator free half strength basal medium. The well rooted plants were hardened successfully in the potting mixture containing coconut husk, charcoal, and brick pieces in the ratio 2:1:1.
Subject(s)
2,4-Dichlorophenoxyacetic Acid/pharmacology , Cell Culture Techniques , Cells, Cultured , Culture Media/pharmacology , Endangered Species , India , Orchidaceae/cytology , Orchidaceae/physiology , Plant Growth Regulators/pharmacology , Plant Roots/drug effects , Plant Roots/physiology , Regeneration/drug effectsABSTRACT
Objective:To study callus induction from different explants (internode, leaf, root) and in vitro plantlets propagation from medicinally important plant Achyranthes aspera L. Methods:Sterilized explants were prepared by using 0.1%HgCl2 and 0.5%Bavistin and callus was obtained when cultured onto Murashige Skoog’s (MS) medium by using different concentrations and combination of 2,4-D, NAA, BAP, IAA, IBA with 3%sucrose and 0.8%agar. Induced callus was immediately transferred to MS medium containing at different concentrations of phytohormones for shootlets and rootlets induction respectively. Results:Sterilization treatment of 0.1%HgCl2 for 2-3 min and Bavistin 0.5%for 10-12 min showed the highest percentage of asepsis and survival rate. Maximum induction of callus was obtained from a combination of 2.0 mg/L 2,4-D and 0.5 mg/L NAA from leaf. Highest shootlets number (4.83±0.17) and length (3.8±0.16) cm were observed on full strength MS medium when fortified with BAP 4.0 mg/L and KIN 0.5 mg/L. Concerted efforts of BAP 2.0 mg/L and NAA 0.5 mg/L on full strength MS medium showed highest leaf number (6.77±0.94). In vitro raised shoots were allowed to root on different strengths of MS medium fortified with IAA and IBA at different concentrations. Experimentally, 3.0 mg/L IBA was enabled to induce maximum rootlets number (10.0±9.82) on full strength MS medium. Afterwards, regenerated shoots with well developed roots were successfully subjected to hardening process and were acclimatized. The survived plantlets showed 66.67%survival frequency without any morphological abnormality. Conclusions: The results demonstrated that different explants were good source of callus induction, morphology analysis as well as indirect plantlets regeneration.
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The present investigation was conducted to study the effects of different concentrations of picloram on somatic embryogenesis induction, development and maturation of three strawberry (Kurdistan, Paros and Camarosa) cultivars. For this purpose, leaf blade, nodal, petiole, stamen and flower bud calli were cultured on MS medium supplemented with picloram at 0.25, 0.5, 1 and 2 mg/L concentrations. The concentration of growth regulator, cultivar and explant type were found critical to somatic embryogenesis induction, development and maturation. Results obtained from the studies revealed that all explants with the exception of petiole and stamen incubated on medium formed embryonic calli. 2 mg/L picloram yielded the highest percentage of embryonic calli and number of globularstage embryos and 1 mg/L picloram yielded the highest number of cotyledonary-stage embryos in all types of explants. The leaf explant calli and Paros cultivar were the most responsive to produce somatic embryogenesis induction, development and maturation.
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A callus induction and plant regeneration protocol was developed from leaf and thorn explants for the plant Ulex europaeus. Explants were incubated on 2 percent sucrose half-strength Murashige and Skoog Medium (MS) with various combinations of plant growth regulators and antioxidants. The best frequency of callus and shoot formation was obtained with 2,4-dichlorophenoxyacetic acid (2,4-D) 1 mg/l x kinetin (Kin) 0.2 mg/l (DK Medium; callus induction) and zeatin (Z) 1 mg/l (DK medium; shoot induction). Both media were supplemented with ascorbic acid 200 mg/l to prevent browning and death of the explants. The regenerated shoots transferred to rooting medium (half-strength MS Medium, 2 percent sucrose) showed rapid growth and development of roots (100 percent). Rooted plantlets were successfully transferred to soil in pots containing a 3:1 mixture of soil and vermiculite.
Subject(s)
Regeneration , Ulex/growth & development , Acclimatization , Plant Shoots/growth & development , Fabaceae/growth & development , GerminationABSTRACT
Objective An effective reproducible protocol for complete plant regeneration via somatic embryogenesis has beendeveloped for Herpetospermum pedunculosum,an endangered Tibetan medicinal herb.Methods The cotyledonexplants used in this study were excised from seedlings germinated in vitro.Callus was induced from cotyledonexplants on Murashige and Skoog's medium,supplemented with 2,4-dichlorophenoxyacetic acid(2,4-D,0.1-1.0mg/L)alone or in combination with 6-benzylaminopurine(BA,0.5,1.0,and 2.0 mg/L).Results The calli showeddifferentiation of globular embryos after three weeks of incubation on MS medium supplemented with variouscombinations of BA and NAA.Sixty-two percent of the embryogenic calli produced somatic embryos in MS basalmedium supplemented with BA(1.0 mg/L)+NAA(2.0 mg/L).The addition of KN(0.5 mg/L)to MS mediumcontaining both BA and NAA(2.0 mg/L each)significantly increased the frequency of somatic embryogenesis.Themaximum percentage of embryogenic calli formation was 83%,and globular embryos formed and germinatedsuccessfully in this medium.Then,transferring the regenerated plants from this medium to hormone-free MSmedium will further enhanced the development of the plants,and the healthy plantlets are formed successfullywithin four weeks.The plantlets were transferred to soil to acclimatize under greenhouse conditions and 75%survived.Conclusion Somatic embryogenesis protocol as reported here can play a key role in the propagation andconservation of this endangered species.
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Objective: The callus induction and subculture condition of Taxus chinensis var. mairei were optimized in the experiment. High Taxol-yielding callus was obtained, and the correlation between expression patterns of Taxol biosynthesis related genes and Taxol content was defined. Methods: Optimal hormone composition and concentration for induction and subculture of callus derived from T. chinensis var. mairei were briefly studied. The growth characteristics and Taxol content of calli derived from T. chinensis var. mairei (including vitro embryos, juvenile stems, and juvenile buds), T. media (vitro embryos) and T. brevifolia (vitro embryos) were compared. Expression of Taxol biosynthesis related genes in the above different calli was analyzed. Results MS Medium supplemented with 3.0 mg/L 2,4-D was suitable for callus induction of T. chinensis var. mairei, the induced rate is up to 92%. Subculture medium supplemented with 2.0 mg/L 2,4-D and 1.0 mg/L 6-BA was suitable for Taxol biosynthesis of T. chinensis var. mairei. Under the same culture conditions, Taxol content of calli derived from vitro embryos of T. chinensis var. mairei was the highest and had reached 0.027% of callus dry weight. The genes of Taxol biosynthesis related enzymes - geranylgeranyl diphosphate synthase (GGPPS), taxadiene synthase (TASY), taxane-10β-hydroxylase (T10βH), 10-deacetylbaccatin III-β-10-O-acetyltransferase (DBAT), phenylalanine aminomutase (PAM), and 3′-N-debenzoyltaxol N-benzoyltransferase (DBTNBT), were expressed at the high level in the high Taxol-producing callus. Conclusion: Vitro embryos can be used as the first choice expiant source to obtain high Taxol-yielding callus. To improve gene expression level of GG-PPS, TASY, T10βH, DBAT, PAM, and DBTNBT would promote Taxol biosynthesis.
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Four commercially grown wheat varieties of Pakistan, namely Inqilab-91, Chakwal-97, Tatara and Manthar were used for this investigation. For callus induction different concentrations of 2,4-Dichlorophenoxyaceticacid (2,4-D) along with 0.1 mg/L of Kinetin were evaluated. For regeneration initially different concentrations of Indole-3-Acetic Acid (IAA) and 6-BenzylAminoPurine (BAP) were tested. Best hormone combinations were further subjected to Kinetin and 6-ã-ã-dimethylallylaminopurine (2iP). For Inqilab-91, Chakwal-97 and Manthar, 3 mg/L of 2,4-D was found optimum, which induced 83.25 percent, 77.75 percent and 95.20 percent of embryogenic calli, respectively. Maximum callus induction (97.18 percent) was observed in Tatara when 2 mg/L of 2,4-D was used. As regard to regeneration, Inqilab-91, Chakwal-97 and Manthar showed maximum regeneration on media containing 0.1 mg/L IAA, 0.4 mg/L Kinetin and 0.5 mg/L 2iP, regenerating 87.25 percent, 81.75 percent and 68.75 percent respectively. For Tatara maximum regeneration of 12.25 percent was obtained on 0.1 mg/L IAA and 2 mg/L of BAP. Presently optimized regeneration method holds promise for facilitating the deployment of agronomical important trait through genetic transformation for the improvement of this important food crop.