Organogenetic Potential Research Articles

In vitro culture responses, callus growth and organogenetic potential of brinjal (Solanum melongena L.) to He-Ne laser irradiation

The present investigation was designed to analyze the influence of Helium-Neon (He-Ne 632.8nm) laser irradiation on defense enzymes, proline content and in vitro responses of callus induction, shoot initiation and on plantlet regeneration potential of brinjal. The seeds of Mattu Gulla (Solanum melongena L.) were irradiated with 20, 25 and 30J/cm2 of He-Ne laser followed by surface sterilization and sprouted on Murashige and Skoog medium without plant growth regulators. The activity of defense enzymes, proline content and the organogenetic potential of hypocotyl, leaf and shoot tip explants were determined from thirty day old seedlings. During seed germination, most of the seedlings showed normal two cotyledons whereas small number of seedlings showed tricotyledonous at 20J/cm2 treatment and no other morphological abnormalities were observed during further growth and development. There was no substantial variation was noted in both β-1,3-glucanase and chitinase activity as well as proline content which proves the He-Ne laser irradiation does not causes any stresses for the plant. The in vitro culture of hypocotyl, leaf and shoot tip explants from laser irradiated seedlings showed differential responses as compared to un-irradiated control. The laser induced enhancement of callus induction, growth rate of callus tissues and shoot tip, percentage of responses of shoot and root initiation, days to shoot and root initiation, shoots formed per callus, number of roots per shoots, length of roots and nuclear DNA content of in vitro raised plants were evaluated. Among the tested laser doses (20, 25 and 30J/cm2), 25J/cm2 showed significant biostimulatory effect over un-irradiated control seedlings. The present observations reveal and endorsed our earlier reports with substantial enhancement of in vitro and ex vitro by He-Ne laser irradiation.

Somatic embryogenesis induction system for cloning an adult Cyphomandra betacea (Cav.) Sendt. (tamarillo)

Somatic embryogenesis is a valuable tool for plant breeding. In recent years, different aspects related to somatic embryogenesis (SE) induction in tamarillo have been studied at our laboratory. In this work, results concerning the establishment of a protocol for cloning an adult tamarillo tree through SE are presented. Attempts to induce SE in tamarillo from various explants directly taken from an adult tree were unsuccessful and only calli with no embryogenic potential were initiated. To overcome the lack of potential of adult tissues for SE, an indirect approach was attempted in which shoots from an adult tree were first established in vitro and then wounded leaves were used for SE induction. A low rate of embryogenic tissue formation was obtained (19.4%), but it was in the range of initiation rates from leaf explants of in vitro cloned plantlets of different tamarillo cultivars (red, orange and yellow) that originated from a single seedling (13.3–54.4%). High variation in SE initiation among juvenile controls could not be explained by different organogenetic potential, as no significant differences in shoot proliferation or rooting ability during micropropagation could be detected. Subcultures of embryogenic lines from the adult tree allowed us to obtain a large amount of embryogenic tissue that, after 8 weeks on a PGR-free medium, gave an average of 111 plants per gram of fresh mass of embryogenic tissue. A RAPD comparative analysis of somatic embryo-derived plantlets and the donor tree confirmed that the plantlets had no variation in the DNA regions amplified by 12 primers. These results open the way for large-scale cloning of elite tamarillo trees through SE.

Differentiation and growth traits associated with acrotony in the apple tree (Malus ×domestica, Rosaceae)

Branching is a key factor in the evolutionary diversification of plants and is a main criterion for plant architecture analysis. Among descriptive features, acrotony is defined as increased vigor of the vegetative proleptic branches (from dormant buds), from the proximal to the distal part of the parent growth unit. I hypothesized that acrotony could be extended to other, usually poorly described, architectural traits. The study was conducted on two architecturally contrasting apple (Malus ×domestica) cultivars, 'Pitchounette' and 'Chantecler.' The proportion and size of various offspring entities were assessed according to their position along the shoot for 2 years after parent shoot growth. Acrotony was characterized by two inverse phenomena: an acropetal decrease in the proportion of latent buds and of laterals that aborted, and an acropetal increase in the proportion of reproductive laterals among growing laterals. Distally located reproductive laterals had more spur leaves and flowers and higher fruit set and flowered earlier than reproductive laterals lower on the parent annual shoot. The results suggest that the length-based criterion used for acrotony should be integrated into a general conceptual framework in which the organogenetic potential of the axillary meristem increases from the proximal to the distal part of the annual shoot, leading to greater branching density, larger offspring, and a greater propensity for flower bud formation over consecutive years.

In vitro regeneration of Trifolium glomeratum

In vitro regeneration of Trifolium glomeratum, a leguminous forage species, was attempted through leaf, petiole, cotyledon, hypocotyl, collar and root explants and two media combinations. Root and collar explants showed no callus induction. Medium with 0.05 mg dm−3 α-naphthaleneacetic acid (NAA) and 0.10 mg dm−3 N6-benzyladenine (BA) was more effective for hypocotyl explant whereas cotyledon and petiole explant were more responsive to 5.0 mg dm−3 NAA and 1.0 mg dm−3 BA. Friable, green calli obtained from petiole explant on this medium showed organogenetic potential. Modified root-inducing medium having 0.21 mg dm−3 indole-3-acetic acid and 2.5 % sucrose was successful for root induction and plantlets were successfully transferred to field after hardening and Rhizobium inoculation.

Efficient plant regeneration in Holarrhena antidysenterica Wall., from shoot segment-derived callus

A procedure has been outlined for plant regeneration of an important medicinal shrub, Holarrhena antidysenterica, through shoot segment-derived callus. Explants used for callus induction were shoot segments derived from 14-d-old axenic plants on Murashige and Skoog (MS) medium supplemented with 15 μM N6-benzyladenine (BA). A white friable type of callus was obtained in 4.52 μM 2,4-dichlorophenoxyacetic acid and 2.32 μM kinetin which did not have the potentiality to regenerate. High-frequency shoot differentiation was achieved on transferring the friable callus to MS medium supplemented with 17.8 μM BA and 8.0 μM naphthaleneacetic acid. The highest percentage of calluses forming shoots (65.06±2.26) was achieved in this medium. The organogenetic potential of the regenerating callus was influenced by the age of the culture. Rooting was achieved on the shoots using MS medium with 25 μM indolebutyric acid. The plantlets were acclimatized and established in soil. The regenerated plants were morphologically uniform and exhibited similar growth characteristics and vegetative morphology to the donor plants.

Induced androgenesis in tomato (Lycopersicon esculentum Mill.) I. Influence of genotype on androgenetic ability : I. Influence of genotype on androgenetic ability.

The androgenetic ability of 85 Lycopersicon esculentum Mill. genotypes was tested. Callus was induced from anthers of 53 lines and hybrids. Regeneration of plants was obtained only from calli of 15 genotypes. The data obtained clearly showed that the genotype affects induced androgenesis in tomato. The in vitro response of anthers from the cultivars Roma, Pearson, San Marzano, Por, Sar, Vigapol, Day, David and Start, containing the ms 1035 gene, which is responsible for male sterility in tomato, confirm the strongly expressed dependence of both callus induction and organogenetic potential on the homozygous or heterozygous state of that gene. A protocol of callus induction, organogenesis and plant regeneration has been developed. More than 6000 regenerants have been obtained. Most of them showed different morphological alterations and variations in chromosome number (n, 2n, 4n). Some are interesting as source material for tomato breeding.

Vegetative multiplication of date palms from in vitro cultured inflorescences: effect of some growth regulator combinations and organogenetic potential of various cultivars

Des fragments d'inflorescences appartenant a differents cultivars femelles de palmier dattier et a un clone mâle selectionne ont ete mis en culture in vitro en presence de cinq combinaisons hormonales differentes. La formation des bourgeons et leur multiplication etaient meilleures sur le milieu Greshoff et Doy en presence de 0,5 mg L-' d'acide naphtylacetique (ANA), 2 mg L-' de 6-benzylaminopurine (BAP) et 1 mg L -1 de 2- isopentyladenine (IPA). Cependant la regeneration de plantules entieres n'a ete obtenue qu'apres reduction des teneurs en cytokinines ou augmentation des teneurs en ANA a 2 mg L -1 en combinaison avec 1 mg L-' d' IPA ou de BAP. Des coupes histologiques realisees a differents moments de culture ont montre que les bourgeons sont inities a partir des petales en multiplication. Des differences entre les cultivars testes ont ete observees en ce qui concerne les pourcentages de reactivite dans differents milieux de culture. Cette technique est tres prometteuse pour la multiplication de l'espece du fait que des centaines de plantes peuvent etre obtenues a partir d'une seule inflorescence et que chaque pied de palmier dattier peut fournir plusieurs inflorescences par an.

Effects of Genotype and Explant duringin vitroResponse to Cadmium Stress and Variation in Protein and Proline Contents in Linseed

Abstract The effects of genotype and explants in influencing in vitro growth in the presence and absence of a selection pressure, viz. the heavy metal cadmium were studied. The response to callus-induction and organogenesis by different explants of three Indian cultivars of linseed ( Linum usitatissimum) viz. Garima, Gaurav, and Kiran was investigated. Of the explants, hypocotyl gave the best in vitro response. Among the cultivars under control conditions (O Cd), Gaurav and Garima fared much better in callusing response compared to Kiran, but Garima and Kiran showed higher organogenetic potential. However, in the presence of Cd (100 μ M ) callusing was not appreciably affected in any of the cultivars, but organogenesis was highly inhibited in both Garima and Gaurav; Kiran appeared to be least affected by the presence of Cd. Genotype, explant, and genotype–explant interactions had highly significant effects on both callusing and organogenesis, but cadmium treatment had a greater effect on organogenesis. To estimate the stress responses due to cadmium treatment, the total protein and proline content in callus lines of the three cultivars was also studied. Both parameters showed an increase due to cadmium, but the genotypic differences in the three cultivars remained unchanged.

Factors influencing Agrobacterium tumefaciens-mediated transformation and regeneration of the safflower cultivar ?centennial?

The effects of co-cultivation conditions on transformation efficiency and direct shoot regeneration from seedling explants of safflower cv. ‘Centennial’ were examined. Agrobacterium tumefaciens strain EHA105/p35SGUSInt was more infective than LBA4404/pBI121 as determined by numbers of sectors expressing β-glucuronidase activity. Compared to nontransformed controls, efficiency of direct shoot regeneration was markedly decreased by co-cultivation with EHA105 and the decrease exacerbated by addition of acetosyringone, indicating that a hypersensitive response to bacterial infection may reduce organogenetic potential. Likewise exposure of co-cultivated explants to kanamycin or geneticin in selective media reduced regeneration efficiency. Addition of 500 mg l-1 carbenicillin slightly increased numbers of regenerating shoots. Tranfformed shoots were obtained only when kanamycin selection was initiated 1 or 2 days after co-cultivation. Presence of transgenes in geneticin-resistant shoots was confirmed using polymerase chain reaction and Southern hybridization assays.

Isatin as an auxin source favoring floral and vegetative shoot regeneration from calli produced by thin layer explants of tomato pedicel.

Thin layer explants taken from the pedicels and peduncles of flowering tomato plants yielded calli with great organogenetic potential. Of the 15 cultivars tested, 7 regenerated roots, shoots and eventually entire fruit-bearing plants. Calli grown on modified Murashige-Skoog medium responded to varied auxins and cytokinins with different morphogenetic patterns. Thus, naphthaleneacetic acid yielded root-producing calli, while the auxin precursor isatin (indole 2,3-dione) caused the production of calli with vegetative and floral shoots, rarely yielding roots. This may be related to isatin's slow, steady conversion to an active auxin (Plant Physiol 41:1485-1488, 1966) in contrast with naphthaleneacetic acid's immediate presentation of a high level of active auxin. The highest incidence of vegetative shoot (100%) and flower (50%) formation was obtained with 10 micromoles isatin and 3 micromoles zeatin. A few of the flowers developed into ripe fruits. The high frequency of induction of vegetative shoots and flowers before roots with isatin suggests its utility in micropropagation from plant tissue cultures.

The relationship between polyploidy and organogenetic potential in embryo- and root-derived tissue cultures of Vicia faba L.

The cell cycle was examined in embryo and root explants of Vicia faba in culture to test whether or not polyploidy and aneuploidy affected organogenetic potential. Nuclear DNA contents and the mitotic index were measured in the 0–1 mm apical segment of primary roots of 5-day old seedlings and at various times following transfer to modified MS in darkness or Chu's N6 medium in an 8 h light/16h dark cycle (N6-MS programme) at 20°C. Mature embryos were dissected and cut longitudinally. Each half was cultured on the N6-MS programme. Root explants grown on MS in darkness developed into callus but there was no subsequent organogenesis. Only on the N6-MS programme were new roots initiated from root-derived callus. Using the N6-MS programme, embryo-derived callus became green and after 3 to 4 months, produced roots and shoots. Approximately 40% of these cultures regenerated plantlets. Polyploidy occurred within 24 h of culture irrespective of both tissue source and culture protocol. Variations in chromosome number from 2n=2x=12 were also routinely observed. Thus, calluses had the ability to initiate roots and shoots regardless of persistent polyploidy and aneuploidy. Compared with the baseline of cell cycle data for roots in vivo, the proportions of cells in the different cell cycle phases remained constant. Thus, in V. faba induction of organogenesis seems more related to culture protocols than to specific changes to the cell cycle. The mitotic index was significantly lower in vitro compared with meristems of intact roots.

Analysis of the organogenetic potential of calli of three Canary Island Lycopersicon esculentum land races

The organogenetic potential from callus of three tomato land races from the Canary Islands adapted to semi-arid environment (‘Salvaje’, ‘Rusa’ and ‘Especial’), and one tomato cultivar (‘Meltine’), were examined. The response of four explant types (cotyledon, shoot apex, hypocotyl and root) to nine PGR regimes (BAP at 1 or 2 or 5 mg/l) + either IAA (0.5 mg/l) or 2,4-D (0.5 or 1 mg/l) were measured. BAP at 5 mg/l+IAA at 0.5 mg/l induced most organogenesis in all the explant types for all genotypes. ‘Salvaje’ has one of the highest organogenetic potentials described in tomato.

Regeneration of calabrian pine from juvenile needles

Cytokinins applied in an agar medium induced adventitious buds on cultured needles from seedling of Pinus brutia Ten. Cytokinins applied as pulses to the explants prior to culture were less effective. Irrespective of the mode of cytokinin application, 8 weeks was the time required to bring about bud formation. Organogenetic potential of the cultured needles decreased with chronological age of the explanted seedlings. The induced buds grew into elongated shoots on culture medium without cytokinins, but the inclusion of activated charcoal (1%) doubled the elongation rate. There was an indication that mixtures of cytokinins were more effective than separate cytokinins in producing buds on explants, but the difference between treatments did not achieve statistical significance. Parenchyma cells in mesophyll layers were evidently the target cells responding to the culture conditions. After a period of activity and division in these cells, meristematic zones developed which later led to formation of bud primordia, and subsequently these primordia developed into well-formed adventitious buds. Subsequent rooting (64%) of shoots was achieved using a combination of two auxins and a low level of cytokinin.

Genetic and physiological influences on differentiation in tissue cultures of a legume, Lathyrus sativus

Callus initiation and subsequent organogenetic potential in cultures of shoot meristems of Lathyrus sativus is under genetic control. This genetic influence is observable not only at the broader intercultivar level, but also between genotypes descended from the same cultivar. However, it is possible to achieve higher levels of morphogenetic response even in recalcitrant genotypes by using the physiologically altered explant. On activation of the dormant lateral bud meristems consequent to decapitation of the apical bud, it is possible to induce organogenesis in the tissues of all genotypes. Callus masses arising from the shoot meristems and leaf expiants of cv. 'LSD-3' also exhibit somatic embryogenesis which results in the development of mature plantlets under culture conditions.

Callus and shoot formation in organ and tissue cultures of Hedera helix L., English ivy

Whole embryos and organs excised from ungerminated embryos and from seedlings of H. helix were cultured to assess their capacity to form callus and regenerate organs. There is a high potential for shoot formation in the explanted cotyledons and in the explanted hypocotyl of the ungerminated embryo. Whole embryos and explanted organs from seedlings produced callus rather than organized structures. In contrast to the excised embryo organs, embryo-derived callus had very low organogenetic potential.

Organogenesis in tissue cultures of certain asclepiads

Summary Embryo and organ differentiation in stem callus tissues derived from 5 members of the fam. Asclepiadaceae have been investigated. Pergularia minor formed embryoids and plant-lets, shoot buds and roots during the initial stages of callus growth but showed a decline in organogenetic potential with continuous subculture. In Asclepias curassavica , embryoids were differentiated at the senescent stage of the callus but these never grew into plantlets under any cultural variables. Dregea, Ibatia and Cryptostegia sp. formed profuse calli but lacked the ability to initiate organs.