Grape (Vitis vinifera L.) seedlings grown in vitro were treated with either 200 mM NaCl or 350 mM mannitol for 7 d. Both salinity and osmotic stress caused significant increase in electrolyte leakage. From the three commonly occurring free polyamines (PA), only conspicuous accumulation of putrescine was found in the NaCl-treated seedlings. Four PA biosynthetic genes encoding arginine decarboxylase (pVvADC), S-adenosylmethionine decarboxylase (pVvSAMDC), spermidine synthase (pVvSPDS) and spermine synthase (pVvSPMS) were successfully isolated. While induction of pVvADC was observed from the 1^st day of salt treatment, pVvSAMDC and pVvSPMS were induced only at late stage of stress. As for expression levels of genes in the mannitol-treated seedling, either temporary (pVvADC at day 1) or late (pVvSPMS at days 5 and 7) induction was observed.

A high frequency adventitious shoot regeneration protocol was developed for henbane (Hyoscyamus niger L.) using thidiazuron (TDZ). Hypocotyl, cotyledon and stem explants were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of N⁶-benzylaminopurine and TDZ. MS medium supplemented with 16 μM TDZ was the most effective for providing 100 % regeneration frequency associated with a 19.53 shoots per hypocotyl explant. Plantlets were rooted on MS medium supplemented with different concentrations of indole-3-butyric acid (IBA) and α-naphthaleneacetic acid. High rooting and survival was achieved using half strength MS medium supplemented with 8 μM IBA.

Genetic engineering of rice (Oryza sativa L. cv. Pusa basmati 1) using synthetic Cry1Ac gene has been achieved by "particle bombardment". Scutellar tissues excised after 5 - 6 d from mature seeds cultured on induction medium were bombarded using gold particles coated with a mixture of Cry1Ac and marker genes on medium with osmoticum. Bombarded tissues were subjected to 30 mg dm^-3 hygromycin selection for two cycles. The selected calli after GUS assay were transferred to shoot regeneration medium. Regenerated shoots were rooted and plantlets (T₀) were grown to full maturity. Polymerase chain reaction (PCR) analysis of T₀ plants using Cry1Ac specific primers revealed the presence of Cry1Ac gene in 65 % plants. Phenotypic assay, β-glucuronidase assay and PCR during T₁ generation revealed the inheritance of the Cry1Ac and marker genes along with the native plant genes.

Two successive cycles of mature embryo-derived callus culture separated by one cycle of sexual reproduction of R₀ regenerated plants were performed using two rice (Oryza sativa L.) cultivars in order to gain information upon the nature of somaclonal variation in this species. Plants regenerated after one cycle of tissue culture exhibited higher variability and lower performances than those of initial cultivar. A second cycle performed using R₁ embryos as explants showed that the cellular component of salt resistance in terms of growth and regenerating abilities selected during the first cycle could be transmitted to the progenies. The extent and the nature of somaclonal variation depended on the identity of R₀ mother plant and culture conditions, somaclonal variation being strongly reduced in some families obtained from salt-treated calli.

The effects of photoperiod on the development of in vitro grown plantlets of yam (Dioscorea alata L.), were investigated. Plantlets were transplanted into pots, acclimatizated until they reached vegetative stages V₁ (3 leaves) or V₂ (8 leaves), and then grown under 12-h or 16-h photoperiod. The formation and development of underground tubers was only induced under 12-h photoperiod. Tuber initiation was not related to the initial vegetative stage of plants, and the tubers were visible at about 18 - 24 d. On the contrary, a 16-h photoperiod inhibited tuber formation and stimulated vine and leaf growth. The total dry matter production and the number of leaves per plant of V₁ stage plants were 50 and 30 % lower respectively, after 44 d under 12-h compared to 16-h photoperiod. These parameters were not influenced by photoperiod in V₂ stage plants. Consequently, the effect of 12-h photoperiod on dry matter of V₁ plants was attributed to a source limitation related to the early initiation of tuberization. The transfer of plants grown under 12-h to 16-h photoperiod stopped tuber growth and starch accumulation. On the other hand, it stimulated the shoots and the roots to grow.

Embryogenesis and plant regeneration was achieved from callus cultures derived from unpollinated ovaries of Psoralea corylifolia L. Callus was initiated from unpollinated ovaries on Murashige and Skoog (MS) medium supplemented with 2.2 µM N ⁶-benzyladenine (BA) and various concentrations of α-naphthaleneacetic acid (NAA (2.7 to 10.7 µM) or 2,4-dichlorophenoxyacetic acid (2,4-D (2.3 to 9 µM) alone or in combination. Highly organized embryogenic callus induction, embryo development, proliferation and maturation were achieved on transfer of callus clumps to MS medium supplemented with NAA (0.27 µM) or 2,4-D (0.23 µM) alone or in combination with BA (2.2 to 8.8 µM). Addition of abscisic acid (ABA) (0.95 to 5.8 µM) to the medium enhanced average numbers of cotyledonary stage embryos, the maximum number (34.6 ± 0.7) being obtained on MS medium containing 0.27 µM NAA, 2.2 µM BA and 3.8 µM ABA. Embryos germinated on MS medium supplemented with BA (0 to 8.8 µM). MS medium containing gibberellic acid (GA₃ (0.29 to 5.8 µM) enhanced embryo germination frequency, the highest frequency (66.7 %) occurring on MS medium containing 2.2 µM BA and 4.3 µM GA₃. Effect of several concentrations (3.0 to 6.0 %) of sucrose or maltose was also observed on germination of embryos. MS medium enriched with maltose supported high frequency of embryo germination.

High efficiency shoot regeneration was achieved through leaflet and cotyledon derived calli in Cassia angustifolia - an important medicinal plant. Dark brown compact callus was induced at the cut ends of the explants on Murashige and Skoog's (MS) medium augmented with 1 µM N⁶-benzyladenine (BA) + 1 µM 2,4-dichlorophenoxyacetic acid (2,4-D). Such callus pieces on transfer to cytokinins (BA or kinetin) supplemented medium differentiated shoots within 10 - 15 d. Of the two cytokinins, 5 µM BA was optimum for eliciting morphogenic response in 83.33 and 70.83 % cultures with an average of 4.16 ± 0.47 and 3.70 ± 0.56 shoots in cotyledon and leaflet derived calli, respectively. The addition of 0.5 µM α-naphthaleneacetic acid (NAA) to MS + 5 µM BA further elevated the maximum average number of shoots to 12.08 ± 1.04 and 5.37 ± 0.52 for cotyledon and leaflet calli, respectively. The excised shoots were transferred to a rooting medium containing either IAA (indole-3-acetic acid), IBA (indole-3-butyric acid) or NAA. Nearly 95 % shoots developed an average of 5.4 ± 0.41 roots on half strength MS medium supplemented with 10 µM IBA.

Efficient plant regeneration system from leaf base segments of wheat (Triticum aestivum L.) was developed. The factors affecting the callus formation and regeneration capacity of leaf segments of two genotypes; Bobwhite and Pavon 76, were investigated. The highest number of somatic embryos (SE) was obtained on Murashige and Skoog medium supplemented with 2 mg dm^-3 2,4-dichlorophenoxyacetic acid + 1 mg dm^-3 naphthalenacetic acid (14.7 SE per segment). Highest frequency of embryogenic callus (96 %) and somatic embryo formation (24.3 SE per segment) were achieved in the first segments. The highest plantlet regeneration was obtained after transfer of embryogenic calli to regeneration medium supplemented with 1 mg dm^-3 kinetin (6.3 plantlets per segment).

The effect of five antibiotics: carbenicillin, chloramphenicol, cefotaxime, kanamycin and hygromycin on the organogenesis from callus cultures of Coryphantha elphantidens (Lem.) Lem. have been studied. Carbenicillin and cefotaxime stimulated shoot regeneration from callus. All antibiotics under study suppressed rooting of in vitro formed shoots. After five sequential subcultures on kanamycin supplemented medium, antibiotic resistant callus was obtained. To study the impact of kanamycin on resistant callus, total protein content was also studied. Selected callus showed a remarkable increase in callus mass. Antibiotic resistant plants have been selected by screening callus pieces on kanamycin supplemented media. Total protein content increased with subsequent subcultures in kanamycin resistant callus. The kanamycin selected shoots withstood the stability test after 2 months on antibiotic free medium. Plants were raised from the callus, which formed roots in 20 mg dm^-3 kanamycin, which was under study.

Structure of callus cells of frost-sensitive and frost-tolerant Solanum species and a frost-tolerant cell line (D20-1), selected from S. tuberosum cv. Desirée callus, was studied. Like frost-tolerant species S. commersonii, cells of the frost-tolerant cell line contained starch grains in their plastids. The cells of this frost-tolerant line also possessed an increased number of microbodies containing protein crystals which suggests the involvement of proteins in frost tolerance but the mechanism may differ from that in frost-tolerant species.

An efficient and rapid regeneration protocol was developed using shoot apices from germinating seedlings of two cultivars of sorghum, SPV-462 and M35-1, as explants. A vertical slit given from the base of each dissected apex enhanced the efficiency of callusing response by two fold. MS medium containing 0.5 mg dm^-3 each of 2,4-D and kinetin was most effective in producing friable and embryogenic calli. Scanning electron microscopy of these calli detected somatic embryogenesis. Calli thus induced gave rise to approximately 42 green shoots per callus in both the genotypes when transferred to regeneration medium containing 1.5 mg dm^-3 kinetin.

A protocol for plant regeneration via somatic embryogenesis was developed in two chickpea (Cicer arietinum L.) cultivars ICCV-10 and Annigeri. Somatic embryos were induced from immature cotyledons on Murashige and Skoog's (MS) medium supplemented with different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), α-naphthaleneacetic acid (NAA) and picloram alone or in combination with 0.5 - 2.0 mg dm^-3 N⁶-benzylaminopurine (BA) or kinetin (KIN). NAA was better for somatic embryo induction compared to other auxins. The well formed, cotyledonary shaped embryos germinated into plantlets with 36.6 % frequency on MS medium supplemented with 2.0 mg dm^-3 BA + 0.5 mg dm^-3 abscisic acid (ABA). The frequency of embryogenesis and plantlet regeneration was higher in cv. ICCV-10 as compared to cv. Annigeri. Regenerated plants were transferred to soil (40 % survival) and grown to maturity. Histological studies of explants at various developmental stages of somatic embryogenesis reveled that somatic embryos developed directly from the cotyledon cells and they were single cell origin.

Direct somatic embryogenesis of Frittilaria meleagris L. was induced using leaf base explants excised from in vitro grown shoots. Somatic embryos occurred at the basal part of leaf explants 4 weeks after culture on a Murashige and Skoog (MS) medium supplemented with various concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) or kinetin (KIN). The highest number of somatic embryos (SEs) were formed (9.74) from leaf explant on MS medium supplemented with 0.1 mg dm^-3 2,4-D after 4 weeks of culture initiation. An initial exposure to a low concentration of KIN in the medium also enhanced SEs induction. Our observations by light and scanning electron microscopy revealed that SEs originate directly from the epidermal and subepidermal layers of leaf explant. The developmental stages of somatic embryogenesis from the first unequal cell division through the meristematic clusters, multi-cellular globular somatic embryos to the fully formed cotyledonary embryos were determined. After 4 weeks on MS medium without plant growth regulators, SEs developed into bulblets.

In order to evaluate effects of γ-rays on adventitious root formation and ginsenoside production, embryogenic calli induced from cotyledon explants of Panax ginseng C.A. Meyer were treated with γ-rays of 0, 10, 30, 50, 70, and 100 Gy. The highest frequency of adventitious root formation of 75 % occurred at γ-irradiation of 30 Gy, which is considered adequate dosage for selecting mutant cell lines. Five mutated adventitious roots (MAR)3-lines out of the propagation of 142 adventitious root lines treated with 30 Gy were selected based a 100-fold increase in proliferation rate compared to control adventitious roots (CAR) and content of the seven major ginsenosides (Rb₁, Rb₂, R_c, R_d, R_e, R_f, and Rg₁) was determined. In the CAR and four of the MAR3-lines (except for MAR3-109), the Rb/Rg ratio was greater than 1.0, thereby indicating altered ginsenoside composition in these root lines. The HPLC analysis of the MAR3-13 and MAR3-26 lines confirmed different ginsenoside profiles, including the three unidentified ginsenoside candidates, Gm₁, Gm₂, and Gm₃. The ginsenosides of the MAR3-13 and MAR3-26 lines showed high hydroxyl and superoxide radical scavenging activities.

An efficient propagation system via somatic embryogenesis and shoot organogenesis and plant regeneration system for endangered species Primulina tabacum Hance was established. Thidiazuron (TDZ) was the key plant growth regulator for inducing somatic embryogenesis and kinetin (KIN) and 6-benzylaminopurine (BAP) were the key cytokinins for inducing shoot organogenesis from leaf explants. TDZ combined with BAP or KIN in the induction Murashige and Skoog medium induced both somatic embryos and adventitious shoots. Leaf explants with abaxial site in contact with the medium induced less somatic embryos or adventitious shoots compared to inversely placed leaf explants and the optimum pH was 6.5-7.0. Secondary somatic embryos or adventitious shoot could be induced from primary somatic embryos using TDZ and BAP. Shoots developed adventitious roots on rooting medium containing 0.5 μM indole-3-butyric acid and 0.2 % activated carbon. Over 90 % of plantlets survived following acclimatization and transfer to potting mixture (sand:Vermiculite:limestone; 1:2:1).

Unfavourable environment brings many kinds of stresses to plants. To survive such stresses, efficient resistance is required for the plants. Multifunctional genes enable the cross-talk among the various abiotic stress resistance systems. This paper reviews the action mechanisms of multifunctional genes. These genes can be classified into three groups: genes encoding diverse proteins through mRNA splicing (e.g. AOX in rice); genes like BADH, P5CS and HAV that control drought, salinity, osmotic and heat stress resistance; and a gene family, for example AQP, controlling transport of many compounds including water and nutrients. These genes participate in signal sensing and transduction, transcriptional regulation and functional gene activation during stress resistance induction. Furthermore, it should be noted that, under abiotic stresses, the regulation cascades are mutually interdependent and there also exists a close correlation between those cascades and normal plant growth and development.

A novel protocol for plant regeneration from cotyledon explants of eggplant (Solanum melongena) reducing concentration of sucrose was established. The most efficient bud induction medium consisted of Murashige and Skoog (MS) medium supplemented with 2.0 mg dm^-3 zeatin, 0.1 mg dm^-3 indoleacetic acid and 10 g dm^-3 sucrose. After 15 d, the shoot buds were fragmented and transferred to the shoot elongation MS supplemented with 1.0-2.0 mg dm^-3 gibberellic acid and 4.0-8.0 mg dm^-3 AgNO₃, which promoted shoots elongation. The genetic stability of the regenerated plants was analyzed by flow cytometry, RAPD and SSR molecular markers. The results indicated that almost no somaclonal variation was detected among the regenerants.

The DIANTHIN gene encoding a ribosome-inactivating protein (RIP) from Dianthus caryophyllus L. was tested for negative selection in tobacco and rice. Tobacco leaf discs and scutellum-derived callus of rice were transformed with Agrobacterium tumefaciens strain LBA4404 (pSB1, pJAS1). pJAS1 harbors the DIANTHIN gene under the control of the CaMV 35S promoter. Tobacco transformation efficiency, in comparison to pCAMBIA1301, was reduced by 87 % in pJAS1-transformed leaf discs. The DIANTHIN gene proved to be completely toxic to tobacco as all the recovered hygromycin-resistant transgenic plants harbored truncated T-DNAs with deletions of the DIANTHIN gene. Transformation of the DIANTHIN gene under a Mungbean yellow mosaic virus (MYMV)-inducible promoter did not cause any toxicity in tobacco as reflected by the recovery of transgenic tobacco plants with the complete DIANTHIN gene. Transformation efficiency of pJAS1 did not decline in rice. Interestingly, all transgenic rice plants harbored the complete DIANTHIN gene and expressed the gene. The T₁ transgenic lines showed reduction of sheath blight symptom in the range of 29 to 42 %. The difference in the sensitivity to DIANTHIN between tobacco and rice provides a new direction to study the mechanisms underlying RIP toxicity in plants.

An efficient micropropagation protocol for annatto (Bixa orellana L.) was achieved using nodal shoot tip explants. Shoot buds were obtained on the Murashige and Skoog (MS) medium supplemented with various concentrations and combinations of indole-3-acetic acid (IAA), N⁶-benzyladenine (BA) and triacontanol (TRIA). Maximum of 213 shoot buds along with 18 primary shoots were produced on MS medium containing 0.05 µM IAA, 8.87 µM BA, and 11.2 µM TRIA. The primary shoots elongated best on MS medium containing 6.66 µM BA and 2.45 µM indole-3-butyric acid (IBA). The regenerated shoots rooted best on MS medium supplemented with 4.9 µM IBA. The in vitro rooted plantlets were hardened and establishment rate under field conditions was 70 to 80 %.

Using in vitro culture, we determined the effect of photoperiod during growth of Chenopodium rubrum mother plants on vegetative and reproductive development of offspring. Photoperiod during flowering induction of mother plants (the first 6 d after the germination) has the key influence on seed germination and offspring growth, while offspring flowering and seed maturation is determined by photoperiod their mothers experienced during, and shortly after, flowering induction. The mechanism can be through changes in seed protein pattern which we found dependent on photoperiod experienced by mother plants.

The activities of sucrose phosphate synthase (SPS), sucrose synthase (SUSY), neutral invertase (NI) and soluble acid invertase (SAI) were measured in callus cultures of four Mexican sugarcane cultivars (Saccharum spp.) with a different capacity to accumulate sucrose in stem parenchyma cells. The results indicated that sucrose accumulation in callus was positively correlated to the activity of SPS and SUSY and negatively to the activity of SAI and NI while SPS explained most of the variation found for sucrose accumulation and NI least.

Susceptibility of C. rubrum to Agrobacterium-mediated transformation was demonstrated by inoculating the petioles of in vitro grown plants with A. rhizogenes strain A4M70GUS. Hairy roots were produced in 8 % of explants. They were isolated and maintained on plant growth regulator-free solid or liquid half-strength Murashige and Skoog medium for two years. Hairy root fresh mass increased 30 - 90 folds when grown in liquid medium, which was superior to solid medium, where most of the hairy roots produced calli. When these calli were grown on medium supplemented with 0.5 mg dm-3 thidiazuron, embryo-like structures were obtained. Transgenic status of long-term callus and hairy root cultures was confirmed by histochemical GUS assay, by PCR specific to the uidA, rolA&B and ags genes and by Southern hybridization.

Sucrose, glucose, fructose, and melibiose in different concentrations and combinations in the induction media influenced the viability of the isolated maize microspores and the formation of multinuclear structures. The induction of multinuclear structures on media containing combination of sucrose, fructose and glucose was lower than on media only with sucrose. In media containing melibiose alone or in combination with sucrose, no induction of multinuclear structures was found, however, microspore viability was improved.

An efficient protocol of direct somatic embryogenesis (without involving intermediate callus) has been developed from stem segments and shoot tips of Capsicum annuum L. Explants were cultured on Murashige and Skoog (MS) medium supplemented with thidiazuron (TDZ). Among the various concentration of TDZ tested, 0.5 μM was proved to be best for induction of somatic embryos. Induction, maturation and germination were achieved on the same medium. The shoots developed from somatic embryos were transferred for rooting to MS medium supplemented with indole-3-butyric acid (IBA). All the regenerated plants with 85 % survival rate were normal with respect to morphology and growth characteristics.

Induction of rooting in the microshoots of Plumbago zeylanica was achieved on halfstrength basal Murashige and Skoog's medium supplemented with 0.25 mg dm^-3 indole-3-butyric acid. Rooting was totally inhibited when the microshoots were cultured in vitro under continuous light, however, maximum percentage of microshoots rooted when incubated in continuous light for 4 weeks before transfer to the rooting media. Peroxidase activity increased markedly during root induction indicating a key role of peroxidase in rooting of microshoots of Plumbago zeylanica in vitro.

In this study, Dendrobium Sonia 17 plantlets were used to induce in vitro flowering. Inflorescences were induced and rooting was inhibited in the half-strength Murashige and Skoog medium containing 20 µM N ⁶-benzyladenine (BA). The medium with high P and low N contents was effective to induce inflorescences while the medium with low P and high N contents was only effective to promote forming of shoots. In addition, the induced in vitro inflorescences were able to multiply and maintain without exhibiting a distinctive vegetative phase. Different morphologies of in vitro flowers such as incomplete flower structures, abnormal and unresupinated in vitro flowers were observed.

This report describes in vitro shoot induction and plant regeneration from mature nodal explants of Vitex trifolia L. on Murashige and Skoog (MS) medium fortified with benzylaminopurine (BAP), kinetin (KN), thidiazuron (TDZ), adenine (ADE), and 2-isopentenyladenine (2-iP) (0.25 - 10.0 μM). Multiple shoots differentiated directly without callus mediation within 3 weeks when explants were cultured on medium supplemented with cytokinins. The maximum number of shoots (9 shoots per explant) was developed on a medium supplemented with 5.0 μM BAP. Shoot cultures was established repeatedly subculturing the original nodal explant on the same medium. Rooting of shoots was achieved on half strength MS medium supplemented with 0.5 μM naphthaleneacetic acid (NAA). Rooted plantlets transferred to pots containing autoclaved soil and vermiculite mixture (1:1) showed 90 % survival when transferred to outdoor.

In six cultivars of rice (Oryza sativa L.), Pusa Basmati 1, Basmati 370, Type III, Pant Dhan 4, CSR 10 and Pokkali, embryogenic callus growth, plant regeneration, and proline and total protein contents were studied under salt stress (on agar solidified media containing 0, 0.5, 1.0, 1.5 and 2.0 % NaCl). Four weeks after inoculation the callus fresh mass decreased with increasing salt concentration in all the six cultivars. The regeneration frequency in salt stressed callus was also lower as compared to control. 15 d and 30 d after inoculation proline content increased several fold whereas total protein content decreased markedly with increase in salt concentration.

Specific activities and isoform patterns of peroxidases, acid phosphatases, DNases and RNases were studied in relation to in vitro rooting of Petunia × hybrida microshoots in the presence of 4 µM indole-3-butyric acid (IBA). Specific activities of the above enzymes increased in the course of rooting. Rhizogenesis could be related with an increased specific activity of peroxidases during the initiation phase, in parallel with increased lignin content. Twelve peroxidases, six anionic (A1-A6) and six cationic (C1-C6), seven acid phosphatases (ACP1-ACP7), seven RNases (R1-R7) and four DNases (D1-D4) isoforms were detected following native PAGE. Variation in the number of the above isoforms and their quantity was observed during different stages of rooting. Particularly, A2, A3, C3, C4, C5, ACP2, R1, R2, R3, and D4 isoforms appeared after the induction phase and could be related to emergence of root primordia. Additionally, R3 and D4 could be associated with cell division and differentiation, since these are only expressed in rooted microshoots. Moreover, the higher number of roots in IBA-treated microshoots could be related to the higher expression of RNase and DNase isoforms during initiation and expression phases.

The induction of defence compounds and enzymes involved in the phenylpropanoid pathway were studied in the ripe and green chilli fruits inoculated with Colletotrichum capsici and Alternaria alternata. Total phenols and the activity of phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenol oxidase (PPO) and catalase (CAT) increased in the inoculated ripe and green chilli fruits compared to the corresponding healthy fruits. Total phenols and the activities of the enzymes were at the maximum 2-3 d after inoculation and thereafter declined sharply in ripe chilli fruits, whereas slowly in green chilli fruits. In comparison with ripe chilli fruits, green chilli fruits were more resistant as they showed higher accumulation of total phenols and also higher activities of enzymes.