Regeneration capacity of mature embryo-derived callus in barley ( Hordeum vulgare L.) (original) (raw)
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Regeneration capacity of mature embryo-derived callus in barley ( Hordeum vulgare L.)
Acta Biologica Hungarica, 2009
In this study, induction of regenerable callus from mature embryos in eight Turkish barley varieties was analysed by using different plant growth regulators (PGRs). Varying concentrations (0.5-4 mg l-1) of 2,4dichlorophenoxyacetic acid (2,4-D) and dicamba (3,6-dichloro-o-anisic acid) were tested for callus induction from mature embryos. Highest percent of callus induction was observed in Bornova 92 variety (98.3%) on MS medium supplemented with 4 mg l-1 dicamba. Calli were transferred to regeneration media with 0.5 mg l-1 dicamba, 0.5 mg l-1 zeatin riboside (ZR) and 2 mg l-1 thidiazuron (TDZ). Low concentrations of dicamba induced multiple shoots during callus regeneration. When the effect of precultivation with 2,4-D or dicamba on the shoot induction were evaluated, lower concentrations (< 4 mg l-1) of auxins have been found optimal. On the regeneration medium with 0.5 mg l-1 dicamba, shoots were able to elongate up to 20 cm and shoot numbers were between 1-23 per callus. The use of ZR led to formation of short shoot buds and somatic embryos in 2 weeks period. The effect of TDZ was different from other PGRs by inducing green solid sectors on calli surfaces (Total 51 sectors/20 callus/Akhisar variety). Five plantlets have been grown from these solid cell clumps and transferred to specific media for root formation. As a result, five varieties (Süleyman Bey, Bornova 92, Vamýk Hoca, Kaya and Akhisar) tested in our study showed the potential to produce regenerable callus by using low amounts of dicamba or TDZ. The optimization process starts from culturing embryos to plantlet formation took nearly 4 weeks.
For understanding the effect of cloned genes and related molecular mechanisms an efficient and reproducible regeneration and stable genetic transformation protocol is necessary. The crucial factors for enhancing successful regeneration are genotypes, explants tissue, combination and concentration of growth regulators, and culture conditions.The present study was undertaken to optimize the concentration of growth regulators for callus induction and subsequent organogenesis in OryzarufipogonL. WRA21.Calli were induced from seeds of WRA21 on MS medium supplemented with 2,4-D (3.0 mg/l), BAP (0.25 mg/l) and proline (0.6 g/l) gelled with gelrite (3.0 g/l), after 10 days of seed culturing. The embryogeniccalli were regenerated under subdued light at 25°C ± 2°C on MS regeneration medium supplemented with BAP (3.0 mg/l), NAA (0.2 mg/l) and gelled with agar (8 g/l) in combination with gelrite (2.0 g/l) and regeneration occurred in 20 days. The regeneration percentage achieved was very high (82.66%). The plantlets were hardened and transferred to soil in pots. The regeneration protocol so developed through embryogeniccalli was highly reproducible. The plants showed normal growth and flowering under greenhouse conditions.
Journal of Plant Physiology, 1999
A reproducible culture system has been established for embryogenic callus formation and plant regeneration from leaf base segments of barley. The influence of genotype and culture conditions was studied. It was found that a high zinc level in the medium used for germination of the donor seeds resulted in an extension of the basic leaf sector from which callus was formed. Dependent upon genotype and culture conditions, plantlet formation was obtained either through somatic embryogenesis from primary soft callus, somatic embryogenesis from compact callus derived from the primary callus, multiple shoot differentiation from green spots of compact callus, or direct somatic embryogenesis without intermediate callus formation. Plant regeneration was obtained from all genotypes tested. Up to 12 plantlets were formed per segment. Since multiple plants can be regenerated by culture of leaf segments derived from seedlings grown in vitro there is no need of producing donor plants in the greenhouse. The system constitutes a promising basis for genetic transformation experiments.
In vitro regeneration protocol development via callus formation from stem explant of tomato
2015
The experiment was conducted on in vitro regeneration of tomato at the Plant Biotechnology Laboratory, Department of Horticulture, Patuakhali Science and Technology University, Patuakhali. The objective was to develop an efficient regeneration protocol in tomato through callus induction for subsequent plantlet regeneration. Seeds were inoculated on MS medium where germination rate was 78.4%. The stems of in vitrocultured seedlings were used as explants. Different concentrations and combinations of growth regulators were added to MS medium to observe their efficacy on callus induction, shoot initiation and root formation. Stem explants cultured on MS medium fortified with 2 mg/L BAPgave the highest number of shoots (3.0) at 45 DAC. Among the concentrations of PGRs, 0.25 mg/L IAA produced the highest length (4.064 cm) of plantlets, number (5.0) of leaves and fresh weight (0.663 g) of plantlets with the stem explants at 45 DAC. The concentration of 0.5 mg/L IAA produced the highest number (21.00) of roots/plantlet, length (7.676 cm) of roots at 45 DAC, from the same explants. The highest survival rate of in vitro regenerated plantlets in the pot was 70.00 % with the stem explants. The results of the current study showed significant increase in the growth of callus of Solanumlycopersicon Mill. Indicating a good efficiency of the optimized media composition and the experimental model used in comparison to other studies of similar nature.
Journal of Plant Biochemistry and Biotechnology, 2011
A protocol for plant regeneration in Indian cultivars of barley (Hordeum vulgare L.) has been developed using mature embryo culture. The influence of various auxins 2,4-D (2,4-dichlorophenoxyacetic acid), Dicamba (3,6-dichloro-o-anisic acid) and Picloram (4amino-3,5,6-trichloropicolinic acid) on the callus induction and subsequent plant regeneration revealed highest percent of callus induction form cultivar (cv) BL 2 on MSB 5 medium (MS salts + B 5 vitamins) supplemented with 6 mg l −1 Picloram, but maximum number of shoot buds (6-13) were regenerated on MSB 5 medium containing 0.5 mg l −1 Picloram. Regenerated shoots were rooted on half-strength MSB 5 medium. Plantlets were successfully transferred to soil and grown to maturity in greenhouse. The effect of copper sulphate revealed significant improvement in callus induction and plant regeneration when the concentration of CuSO 4 was increased to 3 μM (30 times higher than normal MS medium) for cv BL 2. Regeneration potential differed for different cultivars of barley used, with highest for cv BL 2 and lowest for cv BH 924. We conclude that the Indian barley genotypes exhibit plant regeneration from mature embryo cultures. The protocol has potential application in barley improvement through genetic engineering. Keywords Barley (Hordeum vulgare) . Mature embryo . Callus induction . Plant regeneration . MSB 5 medium . Copper sulphate Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid Dicamba 3,6-dichloro-o-anisic acid Picloram 4-amino-3,5,6-trichloropicolinic acid MSB 5 medium MS salts + B 5 vitamins cv cultivar
International Journal of Agriculture and Biology, 2017
Immature embryos are preferred explant source to get regeneration frequency compatible for genetic transformation in wheat. Under natural conditions, immature embryos can be obtained only once a year. Either controlled conditions to grow wheat in off season should be developed or alternative explant should be optimized. To develop mature embryos as explant, two commercial wheat varieties "AARI-11" and "Galaxy-13" were used for the development of regeneration system. Seeds were imbibed with five different concentrations of 2,4-D for 24 h. Embryos were isolated from imbibed seeds, subjected to seven callus induction media (CIM) containing different commercial auxins, and regeneration was achieved on two regeneration media (RM). Effects of genotypes, 2,4-D in imbibition solutions, CIM, and their interactions were studied. Imbibition produced significant effects on callus induction and embryogenesis. Imbibition solution with 8 mg/L 2, 4-D and CIM4 (2,4-D+Dicamba as growth regulator) was found to be most favorable combination for callus induction, and embryogenesis. Both regeneration media responded excellent for regeneration with 81.10% and 80.62% regeneration frequency. Our results also showed that this is not only the regeneration media but also the genotype, CIM and imbibition solutions that played their role for embryogenic callus induction and regeneration. To further improve regeneration the effects of starvation stress, and extended time at CIM were analyzed for embryogenesis induction. The best embryogenesis was observed after 3 weeks of stress and 9 weeks old calli maintained on CIM through sub-culturing.
Euphytica, 1996
Plant regeneration was achieved in Verbascum speciosum Schard. via organogenesis and somatic embryogenesis by culture of mature embryo explants. Two types of calli, embryogenic and non-embryogenic, were induced from mature embryo explants on Murashige and Skoog (MS) medium supplemented with different concentrations of benzyl adenine (BA) and α-naphthalene acetic acid (NAA). In order to further proliferate the somatic embryoids, the yellow and friable embryogenic calli were transferred on MS medium containing 0.5 mg-1 charchol and 0.1 or 1 mg-1 2,4-dichlorophenoxy acetic acid (2,4-D) or into MS medium containing 60 g-1 sucrose, 50 mgl-1 casein hydrolysate (CH), 0.5 mg-1 kinetin (Kin), 5 mg-1 2,4-D and 0.5 mg-1 charchol. Among the 3 tested media, MS medium containing 0.1 mg-1 2,4-D and 0.5 mg-1 charchol was more effective for proliferation of embryonic calli. Somatic embryos were transferred to hormone free MS medium for maturation and shoot regeneration. In addition, shoots and roots regenerated from non-embryogenic calli in hormone free MS medium or containing NAA and BA. Shoot buds were obtained from non-embryogenic calli and they were transferred to MS medium supplemented with 1 mg-1 BA or Kin for further growth and multiplication. Regenerated plants then were potted and maintained in the greenhouse.
JOURNAL OF SCIENTIFIC RESEARCH, 2022
We report on the influence of different plant growth regulators (PGRs) as well as the type of explants on callus mediated regeneration in Turkey berry/pea egg medicinal plant, S.torvum. The explants viz., hypocotyl (0.4-0.8 cm long), cotyledon (0.6-0.8 cm 2) from 3 week old and leaves (1.0 cm 2) from 6 week old in vitro grown seedlings were cultured on MS medium with various concentrations (0.5-4.0 mg/L) of BAP/KIN alone and also in combination with 0.5 mg/L NAA/IAA. The calli developed from all the explants have shown the maximum percentage of response (94% leaf and cotyledon, 88% hypocotyl) and highest number of multiple shoots induction (36.2±1.50 leaf, 33.2±0.25 cotyledon and 23.4±0.11 hypocotyl explants) per explant with maximum length of shoots (3.9±0.18 leaf, 4.0±0.11 cotyledon and 3.7±0.19 hypocotyl) at 0.5 mg/L NAA+2.5 mg/L BAP (leaf), 0.5 mg/L IAA+2.5 mg/L BAP (cotyledon) and 0.5 mg/L IAA+3.0 mg/L BAP (hypocotyl) in comparison to different concentrations of cytokinins used alone and as well as in combination with IAA/NAA. The calli induced from cotyledon explants were found to be more potent in inducing high frequency number of shoots per explant among all other explants tested in the present investigations. Cytokinins BAP/KIN alone or in combination with IAA/NAA was found to be more effective in inducing callus mediated shoot regeneration in all the explants of S. torvum. For in vitro rooting, the elongated microshoots were transferred on to root induction medium (RIM) fortified with 0.25-2.0 mg/L NAA/IAA. Maximum percentage of response (90%), average number of roots (18.2±0.03) per micro-shoot with highest length of roots (7.8±0.09 cms) was observed at 1.0 mg/L IAA. In vitro rooted plantlets were acclimatized in the green house and transferred into field. The survival percentage was found to be 90% and the plantlets were found to be normal in morphology, flowering and fruiting. Thus, the regeneration protocol developed in the present investigation can be used for the conservation and genetic transformation experiments in S. torvum, not only as a medicinal plant but also as a model plant, based on its regeneration potentiality.
The Malnad region located in the Western Ghats of Karnataka is known for the cultivation of indigenous rain fed land race cultivar of rice. The present study was to investigate the callogenic and caulogenic potentialities of the two indigenous rice cultivar namely Karimundaga and Kanadatumba using dehusked mature embryo explants. For callus and shoot bud differentiation, the explants were cultured on Murashige and Skoog (MS) medium supplemented with 2,4-D (1–3 mg/L), IAA (1–2 mg/L), Kn (1–4 mg/L) and BAP (1–4 mg/L). The morphogenic potentialities of the two rice cultivar differed in texture of callus. In both the cultivar callogenic frequency was optimized at 1 mg/L 2,4-D concentration, it was 94% in Karimundaga and 58% in Kanadatumba. Supplementation of IAA either alone (1–2 mg/L) or in combination with Kn or BAP at 1 to 4 mg/L concentration of each induces shoot bud differentiation from the calli. In the cultivar Karimundaga caulogenic frequency was highest (10.60±2.55) at 1.0 mg/L IAA and 4.0 mg/L BAP concentration. While in the cultivar Kanadatumba highest number of shoot buds (7.90±2.69) was differentiated at 1.0 mg/L IAA and 4.0 mg/L Kn concentration. The calli derived regenerants were successfully acclimatized in the greenhouse and agro-morphological variations were evaluated. The growth characteristics and yield related parameters exhibited by in vitro plants were lower than the in vivo plants.
The present investigation was undertaken to determine a suitable media compositions for callus induction and plant regeneration of rice (Oryza sativa L.) through somatic embryogenesis. Murashige and Skoog (MS) media composition and phytohormones (i.e. auxin; 2, 4-D, NAA and cytokinin; BAP, KIN) were tested for high callus induction and plant regeneration. Mature seeds of six elite rice cultivars namely BRRI dhan28, BRRI dhan29, BRRI dhan30, BRRI dhan34, BRRI dhan56 and BRRI dhan57 were used for the experimentation. 2.0 mg/l of 2,4-D + 0.5 mg/l of NAA were found suitable inducing high amount of calli (98.22%) and 2.0 mg/l of BAP+ 1.0 mg/l of NAA + 1.5 mg/l of KIN were found most effective for plant regeneration (80.00%). Among cultivars for callus induction, BRRI dhan29 shows the highest percentage and BRRI dhan30 less and remaining cultivars (BRRI dha28, BRRI dhan34, BRRI dhan56 and BRRI dhan57) show moderate results in the experiments. Among cultivars for regeneration, BRRI dhan29 ...