Combination Of Thidiazuron Research Articles

Prolific regeneration of fertile plants from green nodular callus induced from meristematic tissues in Lathyrus sativus L. (grass pea)

A new in vitro protocol was developed for prolific shoot regeneration from two elite genotypes of Lathyrus sativus L. Vegetative apical and axillary bud explants from greenhouse-grown plants were used to induce regeneration-competent green nodular callus. Callus was induced on two B5 basal media that contained thidiazuron in combination with α-naphthaleneacetic acid or indole-3-acetic acid. Although callus induction was achieved on both media, progression to the formation of green-nodular callus, a distinct morphological feature and prerequisite for regeneration, differed. Shoot elongation was achieved on medium that contained 6-benzylaminopurine, indole-3-acetic acid, and coconut water. Regenerated shoots were readily converted with high efficiency into sexually mature plants either by planting shoots that were induced for rooting on hormone-free B5 basal medium or by grafting of non-rooted shoots.

Regeneration of Fertile Plants from Callus in Phaseolus polyanthus Greenman (Year Bean)

For the first time, plant regeneration of several domesticated genotypes of Phaseolus polyanthus Greenman (year bean) has been achieved. Thidiazuron in combination with indole-3-acetic acid was used to induce morphogenic, green nodular callus from explants that had been obtained either from greenhouse-grown plants or from in vitro -germinated seeds. Of the six genotypes of P. polyanthus tested, five produced shoots in vitro. Regenerated shoots that formed roots in vitro were established in the greenhouse, whereas non-rooted shoots could be established in vitro by grafting. Morphologically normal progeny plants were obtained from the greenhouse-established regenerants. However, by using the same procedure, no regeneration response was observed in two domesticated and two wild genotypes of P. coccineus L. (runner bean). This protocol should help achieve Agrobacterium - or particle bombardment-mediated genetic transformation to improve this important food legume.

Adventitious bud induction in Eucalyptus globulus Labill

Adventitious buds and shoots of Eucalyptus globulus Labill. (Tasmanian Bluegum) have been regenerated from cotyledons and hypocotyls from mature embryos and seedlings. Adventitious buds, were induced at high frequency with 0.05 μM thidiazuron in combination with 0.2 μM 2,4-dichlorophenoxyacetic acid or 5 μM α-naphthaleneacetic acid. Culture of explants in the dark inhibited bud induction, but up to 86% of cotyledons, longitudinally split just prior to culture, produced adventitious buds, in the light. Development of buds into shoots occurred only at low frequency, after transfer to media containing N6-benzylaminopurine.

Differential In vitro Direct Shoot Regeneration Responses in Embryo Axis and Shoot Tip Explants of Faba Bean.

An efficient plant regeneration procedure using embryo axis and shoot tip explants was developed in faba bean (Vicia faba L.). Embryo axes and shoot tips were cultured on MS medium (Murashige and Skoog 1962) supplemented with benzyladenine (BA) and thidiazuron (TDZ). Differential regeneration efficiency was observed due to factors including growth regulator combinations in the medium, explants, basal media and lines used. Explants cultured on medium with TDZ in combination with BA produced a higher number of shoots compared with either cytokinin alone. The embryo axes promoted the formation of more adventitious shoots than shoot tips. The frequency of multiple shoot formation varied among the lines tested. Line 740 cultured on MS medium containing TDZ in combination with BA (2.0 mgl-1 each) gave the highest number of shoots per explant (10.2 and 7.7) from both embryo axes and shoot tips, respectively. Explants cultured on MS or MSB (MS salts+B5 vitamins) produced the highest number of shoots. The efficiency of TDZ combined with BA for multiple shoot induction was not improved when 0.25-1.00 mgl-1 of naphthaleneacetic acid (NAA) was added. Regenerated shoots were rooted on half-strength MS basal medium lacking growth regulators or supplemented with either NAA, IBA or IAA (0.25-1.0 mgl-1) prior to transfer to greenhouse conditions. We developed a rapid, reliable, high-frequency method for producing multiple shoots from faba bean shoot tip and embryo axis explants.

Combined effect of cytokinins on multiple shoot production from cotyledonary node explants of Bauhinia vahlii

Using seedling explants, an improved regeneration protocol was developed for Bauhinia vahlii. A combination of thidiazuron and kinetin (1.0 μM each) increased the number of shoots significantly (p > 0.05) up to four successive subculture cycles. Over 83% shoots rooted on one-fourth strength Murashige and Skoog (MS) medium supplemented with 1.0 μM α-naphthaleneacetic acid (NAA). Fifty percent of plantlets (15 No.) successfully acclimatized in 90 g (w/v) soilrite + sand + soil (2:1:1) in the shed house. Preconditioning at different sucrose concentrations prior to acclimatization showed no effect on percent survival but improved plant quality.

A combination of thidiazuron and benzyladenine promotes multiple shoot production from cotyledonary node explants of faba bean (Vicia faba L.)

A procedure for multiple shoot formation from cotyledonary node explants of faba bean (Vicia faba L.cv.S.Ghdar) cultured on MS medium containing benzyladenine (BA) and thidiazuron (TDZ) was developed. Explants on medium with TDZ in combination with BA produced a higher number of shoots than with either cytokinin alone. The highest number of shoots was obtained when explants from 7-day-old seedlings were cultured on MS medium supplemented with TDZ and BA (2 mgl−1 each) for 31 days before transfer to hormone-free MS medium for elongation. Shoots produced in vitro were rooted on half-strength agar-solidified MS basal medium or with 0.25 or 0.5 mgl−1 naphthalenacetic acid (NAA) prior to transfer to green house conditions. This procedure was found to be applicable to seven other cultivars of faba bean from widely diverse provenances. Thus, it can be advantageously applied to the production of transgenic faba bean plants.

Micropropagation of tea (Camellia sinensis (L.) O. Kuntze) using Thidiazuron

The effect of thidiazuron (TDZ) on the micropropagation of Camellia sinensis (China hybrid) was compared with that of benzylaminopurine (BAP) using nodal segments from in vitro raised seedlings. Extremely low concentrations of TDZ (1pM–100nM) alone were effective in inducing shoot bud proliferation and maintaining high rates of shoot multiplication on hormone-free media. On the other hand, higher concentrations of BAP (1–10μM) and its continued presence were required to initiate and sustain shoot proliferation. While wider ranges of BAP combined favourably with auxins like NAA or IBA, only specific combinations of TDZ and NAA were effective for shoot proliferation. TDZ treated explants yielded healthy shoots, with sturdy leaves, even during the initial stages of growth, whereas, the effect of BAP was cumulative over subcultures in attaining a high proliferative rate.

Combination of thidiazuron and basal media with low salt concentrations increases the frequency of shoot organogenesis in soybeans [Glycine max (L.) Merr.

A successful, efficient system for multiple soybean shoot induction of soybean [Glycine max (L.) Merr.] is reported. Multiple shoots were induced from cotyledonary nodes and hypocotyl segments cultured on media supplemented with 2 mg/l thidiazuron (TDZ) or 1.15 mg/l benzyladenine (BA). It was found that TDZ induced adventitious shoots more efficiently than BA and that hypocotyl segments promoted more adventitious shoots than cotyledonary nodes. The optimal TDZ concentrations for shoot organogenesis from hypocotyl segments were between 1 and 2 mg/l. Basal media also influenced the efficiency of shoot organogenesis. The frequency of adventitious shoot formation tended to increase when the salt concentration in the basal media supplemented with 2 mg/l TDZ was reduced. Two media (1/2B5 and 1/2L2) stimulated shoot organogenesis efficiently from hypocotyl segments. This method can thus be advantageously applied in the production of transgenic soybean plants.

Multiple shoot formation from cotyledonary node segments of Eastern redbud

A procedure for multiple shoot formation from cotyledonary node explants of Eastern redbud (Cercis canadensis L.) cultured on DKW medium containing benzyladenine (BA) and thidiazuron (TDZ) was developed. Explants on medium with TDZ in combination with BA produced higher numbers of shoots than with either cytokinin alone. The highest number of shoots (7.8 to 9.8 shoots per explant) was obtained when explants from 4 to 10 day-old seedlings were treated with a combination of 10 or 15 µM BA and 0.5 or 1.0 µM TDZ for 20 days before being transferred to the same medium without TDZ. The number of shoots formed was increased from 5.8 to 7.2 shoots per explant by cutting through the cotyledonary node prior to culture. Histological studies indicated that the shoots were formed from actively dividing cells located at the axillary bud region. Shoots formed roots in half strength woody plant medium (WPM) supplemented with 10 to 200 µM indole-3-butyric acid (IBA) cultured for 15 days prior to transfer to greenhouse medium.

Regeneration ofSolanum tuberosum cv. Katahdin from leaf expiantsin vitro

A previously published medium has been found to be effective for plant regeneration from leaf expiants ofSolanum tuberosum cv. Katahdin. A number of combinations of thidiazuron and naphthaleneacetic acid, along with a medium previously published for shoot regeneration from protoplast-derived calli ofSolanum phureja, which contained zeatin, indoleacetic acid and gibberellic acid (ZIG), were tested for callus induction, and all calli were transferred to ZIG for shoot induction. We conclude that leaf expiants cultured on ZIG medium at all stages of culture were the most effective at shoot production. A mean of over six shoots were produced per expiant after four months in culture.

Multiple shoot formation from normal and malformed somatic embryo explants of Eastern redbud (Cercis canadensis L.).

A procedure for multiple shoot formation from somatic embryo explants of Eastern redbud (Cercis canadensis L.) cultured on DKW medium containing benzyladenine (BA) and thidiazuron (TDZ) was developed. TDZ in combination with BA produced more shoots than either treatment alone. The highest number of shoots (3.3 to 3.4 shoots per explant) was obtained from partially desiccated and wounded explants treated with a combination of 5 or 10 μM BA and 0.5 or 1.0 μM TDZ for 20 days before being transferred to the same medium without TDZ. The number of shoots formed was increased from 1.5 to 3.2 shoots per explant by cutting through the cotyledonary node prior to culture. In addition, the frequency of explants forming shoots was increased by desiccation of somatic embryo explants to approximately 50% moisture and by using somatic embryos with two well formed cotyledons as explants.

Shoot regeneration from tissue-cultured leaves of the American cranberry (Vaccinium macrocarpon)

A method for shoot regeneration from leaf explants in two cultivars of cranberry (Vaccinium macrocarpon Ait.) is described. Modified Anderson's medium supplemented with combinations of thidiazuron (TDZ) with or without 1 μM NAA (α-naphthaleneacetic acid) was used to optimize shoot regeneration. The effect of light or dark incubation was also determined. Maximum regeneration was obtained in the light in the presence of 10 μM TDZ and 1 μM NAA. While this medium was suitable for leaf explants obtained from shoot cultures, regeneration did not occur from leaves collected from greenhouse-grown plants. Elongation of the regenerated shoot tips did not occur until explants were transferred to growth regulator-free medium at which time only a minority of shoots elongated. Elongated shoots could be dissected away from leaf tissue, rooted easily, and acclimitized to ambient conditions.

Enhancement of ethylene biosynthesis and germination with thidiazuron and some selected auxins in Striga asiatica seeds

Witchweed [Striga asiatica (L.) Kuntze], an economically important parasitic weed on several poaceous crops, is difficult to control. In nature, germination and subsequent morphogenesis of Striga are cued to specific host‐derived chemical signals. Seeds (approximately 2.4 mg) treated with thidiazuron (TDZ) or the auxins 2,4‐dichlorophenoxy‐acetic acid (2,4‐D), 1‐naphthalene acetic acid (NAA), or 2‐(4‐chloro‐o‐tolyloxy) propionic acid (MCPP) produced little ethylene (66‐138 nl l−1). Combinations of TDZ with the auxins increased ethylene production by 4‐ to 18‐fold. Ethylene production was strongly inhibited (86–92%) by aminoethoxyvinylglycine (AVG), inhibitor of 1‐aminocyclopropane‐1‐carboxylic acid (ACC) synthase. Ethylene evolved from seeds treated with TDZ in combination with 2,4‐D increased after a lag period and was promoted by a pretreatment in 2,4‐D. TDZ or any of the auxins, at the rates tested, effected negligible to low levels of germination (0 to 16%), whereas mixtures of TDZ with the above auxins stimulated 38 to 84% germination. Test solutions containing TDZ and indole‐3‐acetic acid (IAA) were, however, less effective. TDZ/auxin‐induced germination was inhibited by AVG and the ethylene action inhibitor silver thiosulfate (STS). The inhibitory effect of the former was reversed by treatment with ACC. In vitro studies revealed negligible germination (< 1%) on control medium. Seeds germinating on media containing TDZ alone developed into seedlings with distinct shoots and rudimentary roots. Seeds germinating on media containing 2,4‐D, irrespective of TDZ concentration, were induced to form calli. The results are consistent with a model in which both germination and subsequent morphogenesis in Striga are associated with exogenous and endogenous phytohormones.

In vitro shoot and floral organogenesis from stamen explants from a Rhododendron PJM group clone

Flower buds were collected from a clone from the Rhododendron PJM group during autumn and winter, disinfested, and dissected. Stamens were placed in vitro on media containing combinations of thidiazuron (TDZ) and isopentenyladenine (2iP) where they produced callus, adventitious shoots, and adventitious floral structures. There were significant interactions between the two cytokinins as well as significant main effects. The best shoot organogenesis was obtained generally when 1 μM TDZ and 25 μM 2iP were in the medium. Thidiazuron was the predominant cytokinin that stimulated the production of adventitious floral structures.

Colletotrichum coccodes and thidiazuron alter specific peroxidase activities in velvetleaf ( Abutilon theophrasti)

The height and biomass of velvetleaf ( Abutilon theophrasti), were inhibited by treatment of young seedlings with the fungal pathogen Colletotrichum coccodes or with the growth regulator thidiazuron (TDZ). Applied simultaneously, these two agents synergistically reduced velvetleaf height and biomass. The use of two different peroxidase substrates, guaiacol and o-dianisidine, detected different peroxidase isoform activities. This study identified C. coccodes- and TDZ-induced changes in peroxidase activity and isoform pattern. Treatment with C. coccodes increased soluble peroxidase activity as measured spectrophotometrically with the peroxidase substrate guaiacol. TDZ treatment had no observed effect on guaiacol-reactive soluble peroxidase activity. Isoelectric focusing analysis of peroxidase isoform activity using o -dianisidine as a substrate demonstrated that both TDZ and C. coccodes caused changes in the temporal expression and activity of specific anionic isoforms. TDZ enhanced the activity of a pI 4·26 isoform while C. coccodes enhanced the activity of a pI 4·03 isoform. The combination of TDZ and C. coccodes caused a simultaneous and rapid stimulation of both these isoforms. Based on these observations, and previously published reports of disease symptomatology, we propose that the increase in guaiacol-reactive soluble peroxidase activity represents a peroxidase-associated defence response, and that the increased activity of the pI 4·03 and 4·26 isoforms is associated with the inhibition of growth. The rapid stimulation of both isoforms following the combined treatment may account for the synergistic inhibition of growth.

Requirement of thidiazuron for healthy protoplast development to efficient tree regeneration of a hybrid poplar ( Populus tremula ß p. alba)

Summary Methods were developed for obtaining high yields of leaf protoplasts of a hybrid clone of white poplar, Populus tremula × P. alba (717-1-B4), as well as rapid and efficient plant regeneration. Protoplasts were isolated from leaves of shoot culture. A precise adaptation of liquid media for preparation and culture of protoplasts led to high rates of cell colony formation, on the average for 30% of initially plated protoplasts. The procedures employed avoided the formation of any exudate. Glucose (9%) as an osmoticum and carbon source was the only sugar leading to high viability. A precise balance of low concentrations of nitrate and ammonium in the culture medium was beneficial for development of healthy cell colonies. A combination of thidiazuron (0.05 µM) as the only cytokinin and 2,4-D (14 µM) as auxin was essential for sustained development of protoplast derived cell colonies. After dilution in liquid medium, about 10% of the colonies formed calli. Plants regenerated through bud formation on practically all protoplast-derived calli. Plants regenerated from 3% of the initially plated protoplasts. In vitro rooting and transfer to soil were also achieved with good efficiency.

Opioids by infusion

To develop a protocol for breaking of seed dormancy and increasing the seed germination rate of Bunium persicum.The seeds were treated with 3.1, 6.3, 12.5, 25, 50 and 100 μmol/L of benzyl aminopurine, gibberellic acid (GA3), thidiazuron (TDZ) and forchlorfenuron. Then, seeds were transferred to two different temperature conditions including room temperature (25 °C) and chilling temperature (2-5 °C).The treatment of moist seeds with chilling temperature (2-5 °C) broke seed dormancy and showed maximum germination, which was 54.7% after 60 d treatment. Also, the treatment of dry seeds with chilling temperature broke seed dormancy with 9.3% germination rate after 120 d. Treatment of seeds with different level of plant growth regulators showed that under moist-room condition, there was evidence of higher and lower seed germination rate: GA3 (100 μmol/L) with 46.7% and TDZ (50 μmol/L) with 6.67% respectively. In addition, the results showed that under moist-chilling condition, TDZ (6.3 μmol/L) with 53.3% seed germination rate had higher influence on breaking seed dormancy. Treatment of seeds with combination of TDZ and GA3 under moist-chilling condition revealed higher rate of breaking of seed dormancy when 6.3 μmol/L TDZ was combined with 100 μmol/L GA3, showing 93.7% germination rate.The effect of plant growth regulators coupled with chilling temperature on breaking of seed dormancy could provide a large number of seedlings while the long juvenile time which is the next restricting factor of plantation still remained. Thus, the subsequent growth of seedlings to provide a large number of corms is necessary for successful plantation.