In vitro regeneration of evergreen azalea from leaves

Rhododendron simsii Hellmut Vogel was regenerated using different types of explants, auxins and cytokinins. After a callus induction phase, with 2,4-dichlorophenoxyacetic acid or -naphthaleneacetic acid, adventitious shoot regeneration was obtained on a medium supplemented with thidiazuron or zeatin. With thidiazuron shoots were small and a subsequent elongation step was required before rooting. An elongation step was not required when zeatin was used. The duration of the callus induction phase was negatively correlated with the regeneration capacity.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - NAA -naphthaleneacetic acid - 2iP 6-(--dimethylallylamino)purine - NOA ß-naphthoxyacetic acid - BA 6-benzyladenine - IBA 1-H-indole-3-butyric acid - IAA 1-H-indole-3-acetic acid - TDZ thidiazuron - WPM woody plant medium     

Somatic embryogenesis and callus production from cotyledon explants of Eastern black walnut

Starting at 8 weeks and continuing until 23 weeks (nut drop) after anthesis,1 m² explants from cotyledons of immature seeds were extracted from Juglans nigra fruits. Explants were placed on Woody Plant Medium with 1 g l^-1 casein hydrolysate and 30 g l^-1 sucrose. The explants remained in light for 4 weeks on primary media containing a 3×3 factorial of 0.05, 0.5, or 5.0 M thidiazuron (TDZ) and 0.1, 1.0, or 10.0 M 2,4-d. Explants were transferred to a secondary medium containing no plant growth regulators and incubated in darkness for 11 weeks. The greatest number of somatic embryos was produced 8, 10, and 12 weeks after anthesis from explants on media with 0.5 or 5.0 M TDZ and 0.1 or 1.0 M 2,4-d. Explants produced the greatest callus volume and dry weight 10, 12, and 14 weeks after anthesis. Throughout the study, callus generally increased with increasing concentrations of both TDZ and 2,4-d.Abbreviations BA 6-benzyladenine - captan 3a,4,7,7a-tetrahydro-2-[(trichloromethyl)thio]-1H-isoindole-1,3(2H)-dione - 2,4-d 2,4-dichlorophenoxyacetic acid - IBA indolebutyric acid - Physan n-alkyl- dimethyl-benzyl ammonium chlorides and n-alkyl-dimethyl-ethylbenzyl ammonium chlorides - TDZ-thidiazuron N-phenyl-N-1,2,3-thiadiazol-5-ylurea     

Micropropagation of Mexican redbud,Cercis canadensis var. mexicana

Mexican redbud (Cercis canadensis var. mexicana) shoot cultures were initiated from explants taken from both mature and juvenile stock plants. Culture conditions affecting shoot growth and proliferation and rooting of three clones were investigated. Shoot growth was best on media supplemented with 0.25% activated charcoal and solidified with 0.2% Gelrite. Four commercially available salt formulations (Anderson's rhododendron medium, WPM, MS, DKW) were tested for growth of shoot cultures, and Anderson's rhododendron basal salt mixture was superior. Axillary shoots grew from explants cultured media supplemented with a wide range of concentrations of benzyladenine and thidiazuron. Benzyladenine at 5.6–22.2 M supported the best combination of shoot quality and number. Rooting of microshoots in vitro was best on half-strength WPM containing 6.71 M naphthaleneacetic acid and 0.1% activated charcoal.Abbreviations BA 6-benzyladenine - IBA indole-3-butyric acid - 2iP 6-(, -dimethylallylamino)purine - DKW Driver & Kuniyuki Walnut - kinetin 6-furfurlaminopurine - MS Murashige & Skoog - NAA -naphthaleneacetic acid - WPM Woody Plant Medium - TDZ thidiazuron - 1-phenyl-3 (1,2,3-thidiazol-5-yl)urea     

Thidiazuron stimulates shoot organogenesis and somatic embryogenesis in white ash (Fraxinus americana L.)

Immature and mature nonstratified seeds of white ash (Fraxinus americana L.) were dissected transversely and 2/3 of each seed was placed onto agar-solidified Murashige and Skoog medium. Adventitious buds, shoots, and somatic embryos formed on callus, cotyledons, and hypocotyls of the resulting seedlings. Shoot organogenesis was induced on explants cultured on medium with 10 M thidiazuron but not on explants on media with benzyladenine (BA) or isopentenyladenine. Not all seed sources were equally capable of shoot organogenesis and embryogenesis. Atypical of adventitious regeneration of other woody plants, mature seed explants of white ash were more organogenic with shoots that elongated better than explants from immature seeds. Somatic embryogenesis was observed in cultures where mature seeds were first cultured for 4 weeks on a medium containing 10 M adenine 2,4-dichlorophenoxyacetic acid in combination with 0.1 and 1.0 M thidiazuron, followed by transfer to a medium containing 0.05 M 6-benzyladenine and 0.5 M naphthaleneacetic acid. Adventitious shoots and epicotyls from both seedlings and germinated somatic embryos were rooted under intermittent mist and acclimatized to the greenhouse.Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - IBA indolebutyric acid - 2iP isopentenyladenine - NAA naphthaleneacetic acid - TDZ thidiazuron-N-phenyl-N-1,2,3-thiadiazol-5-ylurea - WPM woody plant medium     

In vitro regeneration of chickpea (Cicer arietinum L.): Stimulation of direct organogenesis and somatic embryogenesis by thidiazuron

In vitro regeneration in chickpea (Cicer arietinum L.) was achieved by direct culture of mature seeds on Murashige and Skoog (MS) medium supplemented with either N-phenyl-N(-1,2,3-thidiazol-5-yl) urea (thidiazuron, TDZ) or N⁶-benzylaminopurine (BAP). Multiple shoots formed de novo without an intermediary callus phase at the cotyledonary notch region of the seedlings within 2 to 3 weeks of culture initiation. TDZ was found to be more effective compared to BAP as an inductive signal of regeneration. The former induced multiple shoot formation at all the concentrations tested (1 M to 100 M), although, maximum morphogenic response was observed at 10 M concentration. Addition of naphthaleneacetic acid (NAA) alone or in combination with BAP to the MS medium failed to invoke a similar response. When the TDZ supplemented medium was amended with L-proline, the resultant regenerants were mostly somatic embryos. Histological investigations confirmed the switch in the regeneration pathway from directly formed adventitious shoots to embryogenesis. For obtaining plantlets, adventitious shoots were rooted on MS medium supplemented with 2.5 M NAA; somatic embryos were germinated and established on MS medium. Normal plants were regenerated from both adventitious shoots and somatic embryos and transferred to soil.Abbreviations BAP 6-benzylaminopurine - MS Murashige and Skoog [14] basal medium - NAA naphthaleneacetic acid - TDZ thidiazuron [N-phenyl-N(-1,2,3,-thidiazol-5-yl)-urea]     

Adventitious shoot regeneration from leaf tissue of three pear (Pyrus sp.) cultivars in vitro

To develop an adventitious regeneration system for pear cultivars, several experiments were conducted with 2 cultivars of Pyrus communis L. (Seckel and Louise Bonne) and one cultivar of P. bretschneideri Rehd. (Crystal Pear). Half-leaves, taken from shoots proliferating on Lepoivre medium, were plated in petri-dishes on medium supplemented with various combinations of cytokinins and auxins. Cultures of the above cultivars had been established from mature trees. Among the growth regulators tested, thidiazuron (TDZ), combined with naphthalene-acetic acid (NAA), was the most efficient for stimulation of adventitious shoots. The optimum level of TDZ was about 3 uM; shoot regeneration was observed over a wide range of TDZ and NAA concentrations (0.5 to 5 uM and 2.5 to 13 um, respectively). Among different macronutrient compositions, 1/2 and 1/4 Murashige and Skoog were the most effective. Sucrose concentrations (10 to 50 g L-1) had a linear significant effect on shoot regeneration of Crystal Pear.Abbreviations BAP 6-benzylaminopurine - IBA indole-3-butyric acid - NAA a-naphthaleneacetic acid - NoA 2-naphthoxyacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - TDZ thidiazuron (N-phenyl-N'-1,2,3-thidiazol-5-ylurea) - MS Murashige and Skoog (1962) macroelements - L Lepoivre (Quoirin and Lepoivre, 1977) macroelements     

Adventitious shoot regeneration from leaf explants of tissue cultured and greenhouse-grown raspberry

Adventitious shoot regeneration was observed using leaf-petiole explants from shoot-proliferating cultures of Comet red raspberry (Rubus idaeus L.). A maximum regeneration rate of 70% (3.7 shoots/explant) was obtained using 4.5–9.1 M (1–2 mg l^–1) N-phenyl-N-1,2,3-thiadiazol-5-ylurea (thidiazuron or TDZ) with 2.5–4.9 M (0.5–1 mg l^–1) 1H-indole-3-butanoic acid (IBA) or 2.3 M (0.5 mg l^–1) TDZ with 4.9 M (1 mg l^–1) IBA in modified Murashige-Skoog medium. TDZ was more effective than N-(phenylmethyl)-1H-purin-6-amine (BA) at promoting regeneration in combinations tested with IBA (maximum 50% regeneration rate; 1.8 shoots/explant). Variation in the agar concentration or incubation temperature, orientation or scoring of the leaf-petiole explants and use of separate leaf or petiole explants had no effect on shoot regeneration. Incubation in the dark for 1, 2 or 3 weeks prior to growth in the light did not influence the percent regeneration rate but depressed the number of adventitious shoots. Explant source, from micropropagated shoots or greenhouse-grown plants, had an effect on shoot regeneration that was genotype dependent. Only 8 of 22 (36%) raspberry cultivars were capable of regeneration from leaf explants derived from greenhouse-grown plants.     

Adventitious shoot formation from leaf explants of Rhododendron

A protocol for high frequency adventitious shoot regeneration adventitious shoot regeneration from leaf explants of Rhododendron spp. has been developed. The highest percentage of regeneration and the greatest number of shoots were obtained when leaf explants were cultured on Anderson's medium containing 4.9 M IBA and 73.8 M 2iP. Genotypic variation was observed for adventitious shoot regeneration potential among the seven cultivars tested. Regeneration frequencies ranged from 0 to 96%. Lodestar had the highest rate of regeneration after 3 months of culture with 96% shoot regeneration and an average of 14 shoots per explant. Regenerated shoots were rooted in soil in about 2 months. This protocol should be useful in applying gene transfer techniques to Rhododendron improvement.Abbreviations IAA 1-H-indole-3-acetic acid - NAA 1-naphthaleneacetic acid - IBA 1-H-indole-3-butyric acid - 2,4-d 2,4-dichlorophenoxyacetic acid - BA 6-benzyladenine - 2iP N-(3-methyl-2-butenyl)-1-H-purine-6-amine     

In vitro plant regeneration in lateral bud explants of Cryptanthus bromelioides Var. Tricolor M. B. Foster

Lateral buds of Cryptanthus bromelioides var. tricolor variegated ornamental bromeliad, were cultured on Murashige and Skoog's basal medium (MS) supplemented with napthaleneacetic acid, indolebutyric acid, and kinetin, each at 2 mg/l. The basal part of the cultured buds proliferated and produced adventitious shoots. The shoots developed into plants of three different phenotypes: (a) entirely green, (b) striated (green and yellowish-white) and (c) yellowish-white. Rooting could be induced in (a) and (b) types but successful transplantation was possible only with green plants some of which underwent color changes after establishment in soil.Abbreviations BA benzyladenine; 2,4-D, 2,4-dichlorophenoxyacetic acid - IAA indoleacetic acid - IBA indolebutyric acid; 2,i-P, 6 -dimethylallylaminopurine; Kn, kinetin - NAA naphthaleneacetic acid, Z, zeatin     

Adventitious shoot regeneration from in vitro-cultured leaves of Rubus genotypes

Regeneration of adventitious shoots from leaves of several in vitro-cultured Rubus genotypes was affected by media components and incubation conditions. Leaves cultured at 20°C with a photosynthetic photon flux (PPF) of 40 mol m^-2 s^-1 had a higher regeneration frequency and more shoots per regenerating leaf than ones cultured at 25°C with a PPF of either 40 or 80 mol m^-2 s^-1. Thidiazuron (TDZ) was significantly more effective than benzyladenine. Medium containing 1 M TDZ had the highest percentage regeneration for leaves of Autumn Bliss, Canby, Summit and Sentry red raspberries, whereas leaves of MD-ETCE-1 blackberry hybrid responded more to 10 M TDZ. Higher regeneration frequencies were obtained with 0.5 M indolebutyric acid (IBA) than with 1 M, but no significant difference was observed between 0.5 M and no IBA in other experiments. More shoots per regenerating leaf formed on Murashige and Skoog (MS) and N6 media, than on half-strength MS, Anderson, or Woody Plant media for all genotypes tested. The youngest two expanding leaves near the shoot apex were the most regenerative and yielded the highest number of shoots per regenerating leaf.Abbreviations AND Anderson (1980) - BA benzyladenine - IBA indolebutyric acid - MS Murashige & Skoog (1962) - N6 Chu et al. (1975) - NAA naphthaleneacetic acid - PPF photosynthetic photon flux - TDZ thidiazuron (N-phenyl-N'-1, 2, 3-thiadiazol-5-ylurea) - WPM Woody Plant Medium [Lloyd & McCown (1980)]     

Development of adventitious shoots from <Emphasis Type="Italic">in vitro</Emphasis> grown <Emphasis Type="Italic">Cydonia oblonga</Emphasis> leaves as influenced by different cytokinins and treatment duration

The effects of three cytokinins, kinetin 4.5 M (Kin), 6-benzylaminopurine 4.5 M (BA) and N-phenyl-N1,2,3- thiadiazol-5-yl-urea 4.5 M (TDZ), and the effects of different treatment duration on the regeneration of adventitious shoots from quince (Cydonia oblonga Mill.) leaves were studied. In a first experiment, leaves treated with Kin for 0, 8, 16 and 24 d were transferred to BA or TDZ-containing growth medium. In a second experiment TDZ applied for 0, 4, 8, 12, 16 and 24 d was followed by BA. All treatments included 0.5 M -naphthaleneacetic acid (NAA). In the sequence Kin-BA, the production of adventitious shoots decreased and reddish-coloured nodular structures (RNS) of meristematic appearance increased with increasing duration of Kin treatment, while somatic embryo formation was optimal at 8 d. In the Kin-TDZ sequence, shoot production was initially pronounced, but it declined with increasing duration of the Kin treatment, while the number of roots, somatic embryos and RNS increased. TDZ-BA treatments induced marked shoot production, which gradually increased with increasing duration of TDZ treatment. The presence of TDZ and a long treatment duration appeared to be very important factors in inducing caulogenesis. Kin appeared to be effective in shoot induction but not in shoot development; the results of this work demonstrate that RNS were adventitious shoots blocked at an early developmental stage on account of insufficient cytokinin activity. BA was less effective than TDZ in inducing shoot regeneration. Finally, both Kin and BA applied after 2,4-D treatment promoted somatic embryo induction.     

Somatic embryogenesis and plant regeneration from immature embryos of saw palmetto,an important landscape and medicinal plant

Somatic embryogenesis and plant regeneration from immature zygotic embryos was achieved for saw palmetto (Serenoa repens (Bartr.) Small). Embryos, isolated from immature fruit of native-grown plants, were cultured on Murashige and Skoog medium plus 0.15% (w/v) activated charcoal and supplemented with 452 M 2,4-dichlorophenoxyacetic acid (2,4-D) and 14.7 M N⁶-(2-isopentenyl)adenine (2iP). Clusters of somatic embryos developed from all immature zygotic embryos 5 weeks after culture initiation. After 12 weeks, explants were transferred to the same medium with the amount of 2,4-D reduced to 90.4 M which resulted in somatic embryo proliferation. Somatic embryos were then transferred to the basal medium containing 0.9, 9 M thidiazuron (TDZ), or no growth regulator for conversion into plantlets. The 9 M TDZ treatment was ineffective for plant regeneration. However, 12% of the embryos subcultured on 0.9 M TDZ were able to produce complete plantlets. Shoot production was obtained from 35% of the embryos subcultured in the absence of growth regulators. Rooting (100%) was achieved when these shoots were transferred onto medium containing 22.2 M -naphthaleneacetic acid (NAA).     

Rapid plant regeneration of pea using thidiazuron

A simple and rapid pea regeneration procedure was developed. An average of up to 20 shoots formed from hypocotyl explants of cvs. Sugar Ann and Patriot cultured on Murashige and Skoog basal medium supplemented with 0.5 or 1.0 M thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea). Hypocotyls of Puget and Sugar Daddy did not respond. Regenerated shoots rooted rapidly when cultured on Murashige and Skoog basal medium containing either 2.0 M -naphthaleneacetic acid or 1.0–2.0 M indole-3-butyric acid. Seeds were harvested from regenerated plants after only 9–11 weeks.Abbreviations BAP 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy acetic acid - GA₃ gibberellic acid - IBA indole-3-butyric acid - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic acid - TDZ thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea)     

Somatic embryogenesis and shoot regeneration from excised adventitious roots of the rootstock Rosa hybrida L. ‘Moneyway’

Plants were regenerated from excised adventitious roots of the rose rootstock Moneyway via a three step procedure: callus induction, induction of somatic embryos and shoot development. Callus was induced on excised roots after incubation for 4 weeks in the dark on SH-medium (Schenk and Hildebrandt) containing 50 M 2,4-dichlorophenoxyacetic acid. For embryo induction, calluses were transferred to hormone-free SH-medium and incubated for 8 weeks. The use of Gelrite instead of agar during callus induction stimulated somatic embryogenesis (up to 16% of the explants formed organized structures), whereas the presence of 6-benzylaminopurine in this phase inhibited subsequent regeneration. Yellow solid calluses with embryo-like cotyledons or primordia and friable calluses with embryos were selected, and upon incubation in the light shoots developed. Shoot development was faster and more frequent on solid callus than on friable callus (64% and 21% of the calluses finally formed one or more shoots, respectively). Eleven out of thirteen regenerants developed similarly to control shoots. Finally this regeneration method is compared with other systems for somatic embryogenesis and opportunities for the production of transgenic rose rootstocks and rose cultivare are discussed.Abbreviations BAP 6-benzylaminopurine - BM basal medium - BM+ enriched basal medium - 2,4-D 2,4-dichlorophenoxyacetic acid - DAPI-4,6 diamidino-2-phenylindole - FeEDDHA ferric ethylenediamine di(ohydroxyphenylacetate) - FeEDTA ferric ethylenediamine tetraacetate - IBA indole-3-butyric acid     

Indole butyric acid induces regeneration of phenotypically normal apricot (Prunus armeniaca L.) plants from immature embryos

A prerequisite for most transformation systems is an efficient and reliable method to regenerate phenotypically normal plants. Immature embryos or cotyledons were cultured at three developmental stages (stage 1, 2 and 3, PF=3, 30–60, and 100, respectively) from two unrelated apricot genotypes, Zard and NJA82. Explants were cultured on MS media supplemented with either BA or TDZ at four concentrations (0, 0.5, 5.0 or 20 M) and 2,4-D at 0 or 1 M. Stage 1 embryos cultured on MS medium without growth regulators formed embryoid-like structures. Shoot primordia induction was greatest with stage 2 cotyledons on media containing 5–20 M TDZ and 1 M 2,4-D, although shoot morphology was abnormal, especially with the highest level of TDZ. In another factorial experiment, stage 2 cotyledons were cultured on media containing TDZ (0, 5, 7.5, 10, 15 or 20 M) in combination with either no auxin, 1 M 2,4-D, 1 M IBA, or 5 MIBA. Regeneration percentages of 80% or more were observed on media containing 1–5 M IBA and 5–10 M TDZ. The medium containing 5 M IBA and no TDZ exhibited the highest frequency of phenotypically normal plantlet regeneration.Abbreviations BA 6-benzylaminopurine - IBA indole-3-butyric acid - 2,4-D 2,4 dichlorophenoxyacetic acid - 2iP 2-isopentenyladenine - MS Murashige and Skoog basal medium - NAA 1-naphthaleneacetic acid - PF percent fill [(embryo length/seed length) × 100] - TDZ thidiazuron [N-phenyl-N(1,2,3,-thidiazol-5-yl)-urea] - WPM McCown's woody plant medium     

Plantlet regeneration from mesophyll protoplasts ofBetula platyphylla var.japonica

Summary InBetula platyphylla var.japonica, colonies were induced efficiently from mesophyll protoplasts cultured in half strength MS (1/2MS) liquid medium containing 0.6 M mannitol, 0.09M sucrose and 1 M 4-PU and 1 M NAA at a cell density of 5 × 10⁴/ml. The colonies grew actively and developed into callus after 3 months of culture.Roots differentiated from the protoplast-derived white calluses cultured on the 1 /2MS solid media supplemented with 0.1–1 M 4-PU and 1 M NAA, and 10 M zeatin with no supplementation of NAA. Furthermore, the protoplast-derived green callus differentiated shoots with 1/2MS solid medium containing 1 M 4-PU or 10 M zeatin with no supplementation of NAA. When shoots obtained were cultured on the cytokinin-free MS solid medium with 2.5 M IBA and 0.1 M NAA, they rooted and developed into plantlets after one month of culture.The phenylurea-type cytokinin, 4-PU, was effective for plantlet regeneration from the mesophyll protoplasts ofB. platyphylla var.japonica. This suggests that there is potential for the use of 4-PU in the culture of protoplasts in many forest tree species.Abbreviations BAP 6-benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - FDA fluorescein diacetate - IBA indole-3-butyric acid - 2ip N ⁶-(2-isopentenyl)-adenine - NAA 1-naphthaleneacetic acid - 4-PU N-(2-chloro-4-pyridyl)-N–phenylurea - TDZ thidiazuron     

Micropropagation of Withania somnifera from germinating seeds and shoot tips

Shoot multiplication was achieved in vitro from shoot tips of aseptically germinated seedlings of Withania somnifera L. using low concentrations of 6-benzyladenine (BA), viz. 2.2, 4.4 and 8.9 M. Maximum number of shoots were obtained when 2.3 M 2,4-dichlorophenoxyacetic acid (2,4-D) or 2.5 M indolebutyric acid (IBA) was added to medium containing 4.4 M BA during initiation of shoot multiplication, but not when added later. Direct multiple shoot initiation was also obtained from germinating seeds in the presence of BA alone. Rooting was successful in excised shoots grown on growth regulator-free MS medium. Rooted shoots were successfully established in soil in a greenhouse.     

In vitro regeneration through organogenesis of Meconopsis simplicifolia — An endangered ornamental species

Meconopsis simplicifolia (D.Don) Walp. could be propagated by induction of adventitious shoots from callus produced on hypocotyl, cotyledon and rosette leaf explants of 4-month-old seedlings. Callus was initiated on agar-solidified Murashige and Skoog medium supplemented with 1 M kinetin +10 M -naphthaleneacetic acid (NAA). Shoots formed when the callus was subcultured on medium supplemented with kinetin or benzyladenine (BA) in combination with NAA, indoleacetic acid, indolebutyric acid or gibberellic acid. Excised shoots were rooted on medium containing auxin with 10 M NAA producing the best rooting (55%).Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy-acetic acid - FAA formalin-acetic acid-alcohol - GA₃ gibberellic acid - IAA indole-3-acetic acid - IBA -indolebutyric acid - NAA -naphthaleneacetic acid     

Factors effecting adventitious shoot regeneration from leaf explants of quince (Cydonia oblonga)

A procedure for adventitious shoot regeneration from leaf explants of quince (Cydonia oblonga Mill.) using thidiazuron (TDZ) was developed. Excised leaves of cultures grown on Murashige and Skoog (MS) medium containing 5 M benzyladenine (BA) and 0.9% Gibco Phytagar were used. Several experiments were conducted to determine optimum concentrations of thidiazuron, -naphthaleneacetic acid (NAA) and sucrose. When the medium contained 1.5 M TDZ and 2.5 M NAA, 85% of the discs regenerated shoots with an average of eight shoots per leaf disc. An incubation period of three weeks in the dark was necessary for optimum shoot regeneration. Leaves excised from four to six-week-old cultures gave a higher percent shoot regeneration than leaves from cultures older than six weeks. Regeneration percentages were significantly reduced when sucrose concentration in the medium was less than 3%. A significantly higher percentage of shoots regenerated when leaf discs were placed on the regeneration medium abaxial side down as compared to the adaxial side.Regenerated shoots were cultured on MS medium containing 5 M BA and rooted on half-strength MS medium containing 10 M NAA. Rooted plantlets were acclimatized to greenhouse conditions for evaluation of any somaclonal variation. The importance of these findings are discussed in relation to in vitro improvement of plants.Abbreviations BA benzyladenine - MS Murashige & Skoog (1962) salt mixture - NAA -naphthaleneacetic acid - TDZ thidiazuron (N-phenyl-N'-1,2,3,-thiadiazol-5-ylurea) Approved for publication by the Director, West Virginia Agric. and For. Expt. Sta. as Scientific Article No.2346     

Tissue culture and micropropagation of Cuphea ericoides,a potential source of medium-chain fatty acids

Plant rgeneration occurred on leaf-and stem-derived callus of Cuphea ericoides Cham. & Schlechtd obtained in Murashige and Skoog medium supplemented with auxins [indole-3-acetic acid (IAA), -naphthaleneacetic acid (NAA) or 2,4-dichlorophenoxyacetic acid (2,4-d)] plus cytokinins [6-benzyladenine (BA) or kinetin]. These calluses were subcultured and showed vigorous growth. When subcultured on medium containing 2.22 or 4.44 M BA, the calluses showed profuse regeneration of shoots whereas those subcultured on medium supplemented with 2.69 M NAA or 0.226 M 2,4-d produced numerous roots. Isolated shoots rooted on Murashige and Skoog medium lacking growth regulators or containing 0.54 M NAA or 0.49 M indole-3-butyric acid (IBA). Plantlets were acclimatized to greenhouse conditions.Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - MS Murashige & Skoog medium - NAA 1--naphthaleneacetic acid