Endosperm Explants Research Articles

The appearance of amyloplasts as one of the features distinguishing the initial stages of the development of isolated endosperm in kiwiberry

The potency of plant cells’ reaction to dedifferentiation and then proliferation corresponds with cell reorganization. The details of cell modifications may rely on the fate of the cells. In this study, the characteristics of an endosperm and specific storage tissue under organogenic and non-organogenic development pathways were analyzed. The experimental system was based on isolated endosperm explants (with the seed coat and embryo removed) of kiwiberry cv. Bingo, placed on organogenic (OCIM) and non-organogenic (NOCIM) callus induction media that differed in the type and concentration of plant growth regulators. Histological and ultrastructural analyses of freshly isolated endosperm and explants afer selected periods of the culture were performed using stereoscopic, bright field, and transmission electron microscopy techniques. The dedifferentiation for both pathways– organogenic and non-organogenic – was similar at the morphological level for the first week of the culture. The histological and ultrastructural analyses revealed limited amyloplasts in the cells on OCIM. e organization of proliferated tissue differed significantly from two weeks of the culture: it was more compact for OCIM and looser for NOCIM. Additionally, at this period, there were more amyloplasts with starch granules in dedifferentiated cells on OCIM. The limited starch granules in proliferated cells on NOCIM were observed only afer four weeks of the culture. The presence of plastids with starch granules depends on the competence of endosperm-derived callus in kiwiberry to organogenic processes and can be an early marker of callus destination.

Effect of medium composition, genotype and age of explant on the regeneration of hexaploid plants from endosperm culture of tetraploid kiwiberry (Actinidia arguta)

Endosperm, an ephemeral and storage tissue, serves as a source of nutrition and protection during embryo development and germination. It can be used for the cultivation of polyploid plants in vitro. Here, results of plant regeneration and acclimatization from the endosperm-derived calli of four cultivars of Actinidia arguta has been presented. Seeds excised from fresh fruit and dry seeds stored for one year served as the sources of endosperm explants of selected tetraploid cultivars of A. arguta. Callus Induction Medium (CIM; containing 0.25, 0.5, or 1 mg/l of TDZ) and Actinidia Endosperm Medium (AEM; containing 2 mg/l of 2,4-D and 5 mg/l of kinetin) were used to study the organogenic responses of the calli. On AEM, the source of explant did not significantly affect the rate of callus induction for any of the tested cultivars; no organogenic events were observed. In contrast, on CIM both the source of explants and the cultivar origin caused significant differences in callus formation and subsequent organogenic events. Histological and ultrastructural analyses revealed the adventitious nature of shoot bud formation on these media. The most efficient elongation of shoot buds was achieved after transferring organogenic calli with adventitious shoot buds to a medium supplemented with zeatin or meta-topolin. Robust root induction with minimal basal callus formation occurred on the medium with indole-3-acetic acid. Flow cytometric analysis revealed that the nuclear DNA content in the leaves of some regenerants was approximately 50 % higher (4.5 pg/2C) than that in leaves from the tetraploid seedlings (3.1 pg/2C),which confirmed that those regenerants originated from the endosperm. The regeneration of such hexaploid plants was more efficient when endosperm from fresh seeds served as an explant; therefore, fresh rather than dry seeds are recommended for endosperm-derived plant production. The hexaploid plants of A. arguta can serve as an important source of breeding material.

Simultaneous production of triploid and hexaploid plants by endosperm culture with colchicine treatment in diploid Haemanthus albiflos

Haemanthus albiflos (Amaryllidaceae) is a monocotyledonous plant with high ornamental value. To increase the ornamental value of this species, polyploidy breeding was programmed in this study by employing endosperm culture for producing triploid plants. Further, we tried to produce hexaploid plant by colchicine treatment to endosperm-derived callus. As an initial step, endosperm explants harvested 8 weeks after flowering were cultured on Murashige and Skoog (MS) medium containing 5 mg L−1 4-amino-3,5,6-trichloropicolinic acid (picloram) and 5 mg L−1 6-benzylaminopurine (BAP) for callus induction. Induced calli were maintained on the same medium. Somatic embryos and adventitious shoots were observed after transfer the callus segments onto 1/2MS medium without plant growth regulators (PGRs). Further development of somatic embryos and adventitious shoots were achieved by transferring the callus segments onto 0.5 mg L−1 1-naphthaleneacetic acid (NAA) and 1 mg L−1 BAP. Complete plantlets were established after placing the regenerated shoots and developing embryos on 1/2MS medium without PGRs. Flow cytometric analysis revealed that diploid, triploid, and tetraploid plantlets were involved in these regenerants. When the endosperm-derived callus segments were treated with 0.2% (w/v) colchicine for 24, 48, and 72 h, 4 hexaploid plants were regenerated through the same regeneration procedures as mentioned above. Ploidy levels of the representative diploid, triploid, and hexaploid plants were also confirmed by counting chromosome numbers. Availability of endosperm culture in combination with colchicine treatment was discussed as the means for producing polyploid plants. Triploid plants were regenerated from endosperm-derived callus via somatic embryogenesis and adventitious shoot formation in diploid Haemanthus albiflos. Hexaploid plants were induced by colchicine treatment to the endosperm-derived callus.

Callus Formation on Endosperm from Immature and Mature Fruits of Barringtonia Racemosa

Barringtonia racemosa is mangroves type of plant which had been extensively utilized in conventional practices for relieving ailments of pain and inflammation. Many studies have been done on ethnobotanical profiles, pharmacological activities and chemical compounds in Barringtonia racemosa. However, there is a limited study on callogenesis of this plant particularly from different maturity stage of fruits. The present study is to identify the callogenesis of Barringtonia racemosa from endosperm explants of immature and mature fruits in MS medium supplemented with different concentrations of hormones 2,4-Dichlorophenoxyacetic acid (2,4-D) (0, 0.5, 1.0, 1.5 and 2.0 mg/L) and Kinetin (KIN) (0, 0.5, 1.0, 1.5 and 2.0 mg/L). The optimum hormone combination was found in callus grown on endosperm of immature fruits in MS medium supplemented with 1.5 mg/L 2,4-D and 1.0 mg/L KIN. It was also found that the callus in this treatment grew profusely with highest fresh weight (0.513 ± 0.022 g), 100% callus induction and friable callus texture. The callus fresh weight on endosperm explants was higher in immature fruits compared to mature fruits for all the hormone combinations. Therefore, callogenesis were found more efficient from endosperm explant of immature fruits in Barringtonia racemosa species.

Effects of variations in culture media and hormonal treatments upon callus induction potential in endosperm explant of Barringtonia racemosa L.

ObjectiveTo 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. MethodsThe 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. ResultsFrom 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. ConclusionsA reliable protocol for inducing callus formation of profuse and friable morphology in endosperm explant of B. racemosa had therefore been successfully established.

Effects of 2,4-D and Kinetin on Callus Induction of Barringtonia racemosa Leaf and Endosperm Explants in Different Types of Basal Media

Effects of 2,4-D and Kinetin on Callus Induction of Barringtonia racemosa Leaf and Endosperm Explants in Different Types of Basal Media

In Vitro Regeneration of Triploid Plants of Euonymus alatus ‘Compactus’ (Burning Bush) from Endosperm Tissues

Euonymus alatus (Thunb.) Sieb., commonly known as “burning bush,” is an extremely popular landscape plant in the United States as a result of its brilliant showy red leaves in fall. However, E. alatus is also seriously invasive because of its prolific seed production and effective seed dispersal by birds. Thus, development of sterile, non-invasive, seedless triploid E. alatus is in high demand. In this article, we report successful production of triploid E. alatus using endosperm tissues as explants. In our study, ≈50% of immature endosperm explants and 14% of mature endosperm explants formed compact, green calli after culture in the dark for 8 weeks and then under light for 4 weeks on Murashige and Skoog (MS) medium supplemented with 2.2 μM BA and 2.7 μM α-naphthaleneacetic acid (NAA). Approximately 5.6% of the immature endosperm-derived calli and 13.4% of mature endosperm-derived calli initiated shoots within 8 weeks after they were cultured on MS medium with 4.4 μM benzyladenine (BA) and 0.5 μM indole-3-butyric acid (IBA). Eighty-five percent of shoots rooted after culture on woody plant medium (WPM) containing 4.9 μM IBA for 2 weeks and then on hormone-free WPM medium containing 2.0 g·L−1 activated charcoal for 4 weeks. Eight independently regenerated triploid plants have been identified. Triploid plant regeneration rates observed were 0.42% from immature endosperm explants and 0.34% from mature endosperm explants, respectively, based on the number of endosperm explants cultured. Because triploid plants cannot produce viable seeds, and thus are sterile and non-invasive, some triploid E. alatus plant lines reported here can be used to replace the currently used invasive counterparts. Chemical names used: benzyladenine (BA), indole-3-butyric acid (IBA), and α-naphthaleneacetic acid (NAA).

EFFECT OF PHYSIOLOGICAL AGE AND GROWTH REGULATORS ON CALLUS BROWNING OF COCONUT ENDOSPERM CULTURE IN VITRO

The possibility of physiological age and growth regulators affecting callus browning of coconut endosperm was investigated. Solid endosperm explants of four coconut fruits from same brunches of two coconut cultivars “Samoan Dwarf” were grown on modified Murashige and Skoog (MS) formula with addition of 10 mg l-1 putresine, 2.50 g l-1 activated charcoal (AC), 1.70 g l-1 phytagel, 0, 10-6, 10-5, 10-4, 10-3 M 2,4-dichlorophenoxyacetic acid (2,4-D) or 4-amino-3,5,6-trichloropicolinic acid (Picloram)  combined with 10-5M 6-benzylaminopurine (BA). Callogenesis occurred on 98.83% of explants.  Callus browning between different physiological ages (antipodal and micropylar tissues) of coconut endosperm at  9, 26 and 31 weeks of culture (WOC) was significantly different, but not at 16 and 21 WOC. Auxins of 2,4-D and Picloram did not affect significantly callus browning of endosperm cultures. Auxin doses at 10-6, 10-5, and 10-4M decreased significantly callus browning at 9 and 16 WOC, respectively,  but at 10-6M  browning was less significant compared to  other doses at  21 WOC. Auxin dose at 10-3 M caused  less significant  browning compared to  other doses at  31 WOC. The addition of BA decreased significantly callus browning at  9 WOC , but did not affect callus browning thereafter. Key words: Coconut, in vitro, Picloram, 2,4-D, BA Â

The nitrogen-induced recovery of alpha-zein gene expression in in vitro cultured opaque2 maize endosperms depends on the genetic background.

The effect of nitrogen nutrition on the accumulation of seed storage proteins has been studied in vitro by cultivating on agar media maize (Zea mays L.) endosperm explants from seeds at 10 days after pollination. The experiments were performed on various genetic backgrounds bearing different opaque2 (o2) mutant alleles and on the corresponding wild-type lines. In the seed of the o2 genotypes the high molecular weight alpha-zein polypeptides (zHs), whose transcription is Opaque2 (O2) regulated, are absent or extremely reduced. The endosperms were incubated on basal agar medium with amino acid supply. In these growth conditions, fresh and dry weights increased in both wild-type and o2 endosperms, irrespective of the genetic background. In 4 out of the 5 o2 mutant genotypes analysed we detected an accumulation of the zHs similar to the corresponding wild-type explants or seeds. However, in one of these mutants, Mo17o2R, the addition of amino acids to the culture media had no effect on the zH accumulation. We showed that the Mo17o2R behaviour is not due to a negative regulation but to the absence of putative transcription factor(s) able to regulate the zH transcription occurring in the other o2 mutants.

Shoot regeneration from mature endosperm of Passiflora foetida

Murashige and Skoog (1962) medium supplemented with 2 μM 6-benzylaminopurine (BA) induced adventitious shoots on mature endosperm explants, whilst gibberellic acid (GA3) and casein hydrolysate stimulated growth and development of these shoot primordia. Plantlets were successfully weaned in vivo. These plants were found to be triploid and flowered, although fruit set was not observed.

Growth and metabolism of Ricinus communis endosperm in tissue culture

Endosperm tissue from both developing and germinating castor oil seeds (Ricinus communis L.) was grown as a callus in tissue culture. Callus growths were established from endosperm explants (without embryo) at all stages of seed development except from the quiescent unimbibed seed. Best tissue culture growth was observed with endosperm tissue obtained from seeds that had been germinated for 2 days.The lipid reserves diminished in all endosperm tissue which had been established on culture from germinating seeds but the rate of breakdown was much slower in cultures from 0-day germinated seeds. Glyoxylate cycle activity, as tested by acetate-1- and -2-14C incorporation, was not evident in long-term cultures.The process of lipid accumulation found in normal developing endosperm (24 to 36 days after fertilization) was not retained in culture but was replaced by a process resulting in the rapid loss of the already accumulated oil reserves.As far as could be determined the in vivo biochemical capabilities of the castor bean endosperm were not retained for a lengthy period in the tissue culture conditions used.

HEREDITARY CONTROL OF CALLUS FORMATION IN MAIZE ENDOSPERM CULTURED IN VITRO

1. The growth response of maize endosperm excised from immature kernels was studied in vitro, using White's medium supplemented with yeast extract. The endosperm tissue excised 9 days post-pollination generally gave best growth.2. Of four inbred strains (two of starchy, one of waxy, and one of sugary type endosperm) of maize tested, callus tissue was formed only in the endosperm explants from the starchy strains (S41 and S42). The excised endosperm tissues of the F1 kernels obtained by the reciprocal crosses between S41 and S42 showed a better growth in vitro than those of either of the inbred parents. Histological observations of the endosperm in vivo failed to reveal any developmental or cytological differences between the F1 endosperm and the endosperms of the parental strains.3. Among the callus tissues obtained, only those from the strain S42 and S41×S42 have been cultured successfully for three years, while other cultures ceased to grow within a few months.4. Progeny tests of the S41×S42 hybrid suggested that the two strains could be different in two or more genetic factors concerning the callus formation in the endosperm explant, although the phenotypic expression is influenced by environmental and nutritional factors. Furthermore, the genetic study of the waxy strain S43 suggested that it possesses a genetic factor for inhibiting callus formation.5. None of indole-3-acetic acid, kinetin, and crude extracts of established tissue cultures gave a growth promoting effect on excised endosperm tissues, whereas the amino acid components of yeast extract were found to be essential for the callus formation in the explants.