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Eggplant (Solanum melongena L.): tissue culture, genetic transformation and use as an alternative model plant

Eggplant (Solanum melongena L.): tissue culture, genetic transformation and use as an alternative model plant

ABSTRACT – (Eggplant (Solanum melongena L.): tissue culture, genetic transformation and use as an alternative model plant). Eggplant is an agronomically important non-tuberous solanaceous crop grown primarily for its large oval fruit. In popular medicine, eggplant is indicated for the treatment of several diseases, including diabetes, arthritis, asthma and bronchitis. Eggplant is susceptible to a number of diseases and pests capable of causing serious crop losses. This problem has been addressed by hybridizing eggplant with wild resistant Solanum species, which present a wide genetic diversity and are source of useful agronomic traits. The application of in vitro methodologies to eggplant has resulted in considerable success. Eggplant tissues present a high morphogenetic potential that is useful for developmental studies as well as for establishing biotechnological approaches to produce improved varieties, such as embryo rescue, in vitro selection, somatic hybridization and genetic transformation. Taken together, these characteristics also make eggplant a complete model for studies on different areas of plant science, including control of gene expression and assessment of genetic stability of somaclones derived from different morphogenetic processes. In the present study, important factors that affect the efficiency of in vitro regeneration through organogenesis and embryogenesis as well as genetic transformation are analyzed. The potential of this species as a model plant for studying various aspects of plant genetics and physiology is also discussed.
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Phytosteroids from tissue culture of Allium cepa L. and Trachyspermum ammi S prague.

Phytosteroids from tissue culture of Allium cepa L. and Trachyspermum ammi S prague.

Production of secondary metabolites by cultured cells provides a particularly important benefit to manipulate and improve the production of desired compounds; thus biotechnological approaches to increase the concentrations of the metabolites are discussed. Present study deals with the production, isolation and identification of phytosterols from tissue culture of Allium cepa and from plant parts and tissue culture of Trachyspermum ammi. Steroidal analysis of plant parts showed the maximum amount of stigmasterol (0.240 mg/gdw) which was comparatively little less than that of the amount of β- sitosterol (0.295 mg/gdw) in the seeds of T. ammi . The maximum amount of stigmasterol was present in four weeks old tissue of T. ammi (0.249 mg/gdw) whereas the highest content of β- sitosterol was observed in six weeks old tissue of A. cepa (0.315 mg/gdw) However, lanosterol, was present only in the tissue of A. cepa which was maximum in six weeks old tissue (0.039 mg/gdw)
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Tissue culture efficiency of wheat species with different genomic formulas

Tissue culture efficiency of wheat species with different genomic formulas

Mature wheat embryos are thought to be more recalcitrant to tissue culture than immature embryos, due to differences in the physiological and biochemical tissue status (Özgen et al. 1998, Filippov et al. 2006, Delporte et al. 2014, Yang et al. 2015). The acquisition of embryogenic competence in tissues of zygotic wheat embryos was shown to depend significantly on the endogenous level of growth regulators and total contents of phenols and soluble sugars (Jiménez and Bangerth 2001, Yang et al. 2015). With regard to the dormancy of the mature embryos at the stage of culture initiation, the particular interaction of the medium and other environmental parameters with genotype-specific endogenous factors was frequently required to achieve efficient morphogenesis, especially in endosperm-supported culture (Filippov et al. 2006, Bi et al. 2007, Chauhan et al. 2007, Yu et al. 2008, Miroshnichenko et al. 2011, Yin et al. 2011, Chang et al. 2012, Delporte et al. 2014). With the media tested here however, immature embryos are the preferable explant material. Regardless of the low culture efficiency of the studied germplasms, the ability of mature embryo- derived cultures to generate entire green plants ought to be more deeply investigated, along with immature embryo and anther cultures. Currently, albinism still significantly delays breeding programs using doubled haploid technologies, so some comparative investigations may help discover genetic and developmental mechanisms responsible for the reduction of albino plant formation.
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Standardized production of Phyllanthus tenellus Roxb. by plant tissue culture

Standardized production of Phyllanthus tenellus Roxb. by plant tissue culture

P. tenellus is a widely medicinal herb used in Brazil. The large number of species of Phyllanthus genus and their similarity is a disturbed and a risk in folk medicine. As result of the present study, in vitro systems permit the standardized propagation of P. tenellus raw material under controlled and aseptic conditions showing the relevance of this study together with anatomical pattern of this species as a certificate of safe and quality. Furthermore, the establishment of different tissue culture techniques reveals other alternatives to acquire therapeutic metabolites from P. tenellus under aseptic and controlled conditions.
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Ontogenetically-regulated male sterility in tissue culture - induced and spontaneous sorghum mutants

Ontogenetically-regulated male sterility in tissue culture - induced and spontaneous sorghum mutants

Variability of male fertility expression in the AS-1 line, a somaclonal variant obtained from tissue culture of CMS-plant, and in the progeny of revertant ‘124-1’ obtained from fertile tiller, which developed on CMS-plant transferred from the field to the greenhouse, was investi- gated. Both revertants were characterized by similar expression of male fertility during plant ontogenesis: the panicle on the main tiller was al- most completely sterile whereas formation of fertile pollen grains and seed set were observed on the panicles of the shoot tillers. A clear basipetal gradient of male fertility was manifested on all panicles: the base had significantly higher per cent of fertile pollen grains in compari- son with the middle part, while in the top the anthers were either absent or had few sterile pollen grains. Such an ontogenetically-regulated restora- tion of male fertility was controlled by nuclear genes and could be trans- ferred through the pollen in crosses with progenitor CMS-line. Growing of AS-1 plants in the growth chambers simultaneously under a long (16/8) and a short (12/12) daylength conditions demonstrated that differences of fertility level in different tillers was not caused by change of photoperiod during plant ontogenesis and functioning of photoperiod-sensitive fertility restoring gene. Whereas, the ontogenetically-regulated expression of male fertility in both revertants was temperature-dependent and was clearly manifested under relatively cool conditions during 2-week period before _______________________________
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Dissecting mitosis by RNAi in Drosophila tissue culture cells

Dissecting mitosis by RNAi in Drosophila tissue culture cells

However, interpretation of results after RNAi in Drosophila tissue culture cells can be complex since the penetrance of RNAi is not absolute, these cells are hard to synchronize, live cell analysis is not yet well developed and the resulting phenotypes can often be heterogeneous. Here we describe a detailed methodology for the careful phenotypic interpretation after specific gene inactivation by RNAi in Drosophila tissue culture cells, with particular emphasis on one gene involved in mitosis. The gene in cause, encodes for a conserved microtubule-associated protein called MAST/Orbit that, during mitosis, is localized to the mitotic spindle, centrosomes and kinetochores, ending up accumulating in the central-spindle region and ultimately concentrating at the midbody (23-24). Mutations in mast show severe mitotic abnormalities including the formation of mono- and multi-polar spindles organized by clusters of centrosomes (23). The use of RNAi in Drosophila tissue culture cells was of great help in the dissection of the mitotic role of MAST/Orbit, namely its unexpected role in the behavior of kinetochore microtubules (17, 25).
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Tissue culture techniques in the proliferation of shoots and roots of Calendula officinalis

Tissue culture techniques in the proliferation of shoots and roots of Calendula officinalis

Tissue culture of C. officinalis was reproducible for six subcultures, with slight changes in shoot height (4.2-5.4 cm). Plantlets cultured in MS½N produced a maximum of two shoots per explant (Figure 1 and Table 3). Cytokinins improved the production of C. officinalis plantlets when compared to control. TDZ was less effective compared to BAP because shoots from any TDZ concentration failed to elongate (Figure 2), confirming studies where TDZ inhibits shoot elongation (LEDBETTER; PREECE, 2004). Plantlets from control medium and medium supplemented with BAP reached a height of 4.5 cm and 3.3 cm, respectively (Table 3). Forty-five-day-old plantlets developed in TDZ media were transferred to a medium without growth regulators to elongate. Results such as reduced height and vitrification were verified for plantlets cultured with additional TDZ. Four subcultures intercalated by four transfers were required for the TDZ treatments, all within the one-year period. After subculture in 0.8 mg L -1 TDZ, the greatest
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Organotypic tissue culture of adult rodent retina followed by particle-mediated acute gene transfer in vitro.

Organotypic tissue culture of adult rodent retina followed by particle-mediated acute gene transfer in vitro.

Methodology/Principal Findings: We established an interphase organotypic tissue culture for adult rat retinas (.P35 of age) which was optimized from that used for adult rabbit retinas. We implemented three optimizations: a greater volume of Ames’ medium (.26 mL) per retina, a higher speed (constant 55 rpm) of agitation by rotary shaker, and a greater concentration (10%) of horse serum in the medium. We also successfully applied this method to adult mouse retina (.P35 of age). The organotypic tissue culture allowed us to keep adult rodent retina morphologically and structurally intact for at least 4 days. However, mouse retinas showed less viability after 4-day culture. Electrophysiologically, ganglion cells in cultured rat retina were able to generate action potentials, but exhibited less reliable light responses. After transfection of EGFP plasmids by particle-mediated acute gene transfer, we observed EGFP-expressing retinal ganglion cells as early as 1 day of culture. We also introduced polarized-targeting fusion proteins such as PSD95-GFP and melanopsin-EYFP (hOPN4- EYFP) into rat retinal ganglion cells. These fusion proteins were successfully transferred into appropriate locations on individual retinal neurons.
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Air exposure induced characteristics of dry eye in conjunctival tissue culture.

Air exposure induced characteristics of dry eye in conjunctival tissue culture.

There are several animal models illustrating dry eye pathophysiology. Current study would like to establish an ex vivo tissue culture model for characterizing dry eye. Human conjunctival explants were cultured under airlift or submerged conditions for up to 2 weeks, and only airlifted conjunctival cultures underwent increased epithelial stratification. Starting on day 4, the suprabasal cells displayed decreased K19 expression whereas K10 keratin became evident in airlift group. Pax6 nuclear expression attenuated already at 2 days, while its perinuclear and cytoplasmic expression gradually increased. MUC5AC and MUC19 expression dramatically decreased whereas the full thickness MUC4 and MUC16 expression pattern disappeared soon after initiating the airlift condition. Real time PCR showed K16, K10 and MUC16 gene up-regulated while K19, MUC5AC, MUC19 and MUC4 down-regulated on day 8 and day 14. On day 2 was the appearance of apoptotic epithelial and stromal cells appeared. The Wnt signaling pathway was transiently activated from day 2 to day 10. The inflammatory mediators IL- 1b, TNF-a, and MMP-9 were detected in the conditioned media after 6 to 8 days. In conclusion, airlifted conjunctival tissue cultures demonstrated Wnt signaling pathway activation, coupled with squamous metaplasia, mucin pattern alteration, apoptosis and upregulation of proinflammatory cytokine expression. These changes mimic the pathohistological alterations described in dry eye. This correspondence suggests that insight into the pathophysiology of dry eye may be aided through the use of airlifted conjunctival tissue cultures.
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Deep tissue culture and hemoculture in dogs with wounds and sepsis

Deep tissue culture and hemoculture in dogs with wounds and sepsis

Arias et al. (2008, 2011), studying traumatic wounds in dogs, found Pseudomonas spp., Proteus spp. and Sta- phylococcus spp. as the most frequently isolated bacteria. However, in these two studies, the wounds were sampled using swabs, which frequently identify the contaminant mi- croorganisms on the wound surface, such as Staphylococ- cus spp. (Pallua et al. 1999). Similarly, Mouro et al. (2010) used swabs to collect sample from wounds during initial presentation, resulting in the isolation of mainly bacteria belonging to the cutaneous flora. The culture from the ma- terial collected by biopsy in the present study allowed for identifying bacteria not belonging to the normal skin mi- crobiota, emphasizing the importance of differentiating be- tween pathogens responsible for tissue infection and the skin-colonizing agents, which usually do not require treat- ment with antimicrobials and do not interfere with the hea- ling duration (Buchanan et al. 1986, Amalsadvala & Swaim 2006, Meyers et al. 2008, Dryden 2010).
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Building bridges toward invasion: tumor promoter treatment induces a novel protein kinase C-dependent phenotype in MCF10A mammary cell acini.

Building bridges toward invasion: tumor promoter treatment induces a novel protein kinase C-dependent phenotype in MCF10A mammary cell acini.

The complex action of TPA raises the question of which effect or effects are critical for helping cells advance along the pathway of carcinogenesis. Identifying the important events that occur during early stages of carcinogenesis can aid the development of targeted strategies for preventing cancer. To address this question, we investigated the action of TPA in a three-dimensional (3D) cell culture system that uses human cells to model the cellular organization, signaling, and growth constraints of epithelial tissues [12]. Investigating the action of TPA in a 3D human cell culture model could reveal information about the roles of PKC in carcinogenesis that may have been missed by studies conducted in traditional monolayer tissue culture models and in vivo rodent models. We chose the 3D MCF10A human breast epithelial cell system because it recreates important features of in vivo epithelial tissue that affect cell signaling, including the spatial organization of cells, cell polarization, and establishment of a basement membrane [12]. MCF10A cells are immortalized, but nontumorigenic [13]. When grown within 3D culture conditions, MCF10A cells form hollow, spheroid structures referred to as acini. The correct formation of acini requires the tight spatiotemporal regulation of cell proliferation, cell polarization, apoptosis, and growth arrest [12]. The 3D MCF10A model has provided insight into how the expression of different oncogenes disrupts the coordination of these basic cellular functions resulting in changes in the morphology of 3D MCF10A structures that correspond to different stages of carcinogenesis [12]. Altogether, these studies suggested that the 3D MCF10A model could provide an integrated picture of the complex action of TPA, and indicate which effects are the most relevant for carcinogenesis.
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In Vitro Biofilm Formation by Uropathogenic Bacteria and their Antibiotic Susceptibility Pattern

In Vitro Biofilm Formation by Uropathogenic Bacteria and their Antibiotic Susceptibility Pattern

This prospective analytical study was carried out for six months in the department of microbiology after obtaining institutional ethical committee clearance and informed written consent from patients. Total 168 isolates were collected from patients admitted in our hospital with symptoms of UTI for at least two days. Patients of all age groups and of both sexes were included in the present study. Midstream urine samples were obtained after proper anogenital toilet. Samples were inoculated in blood agar and Mac Conkey's agar with calibrated loop to determine Colony Forming Units (CFU). Patients with significant bacteriuria were included in the present study. Organisms were identified on the basis of their growth characters, gram staining and biochemical tests as per the standard recommended procedures [5]. AST was carried out by Kirby Bauer disc diffusion method on Muller Hinton Agar as per Clinical and Laboratory Standard Institute guidelines [6]. Detection of biofilm production was done by using three methods, namely Tube Adherence Method (TAM), Congo Red Agar Method (CRAM) and by Tissue Culture Platte Method (TCPM).
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Bauhinia forficata Link Shoot Regeneration: Histological Analysis of Organogenesis Pathway

Bauhinia forficata Link Shoot Regeneration: Histological Analysis of Organogenesis Pathway

Bauhinia forficata, a leguminous tree, is native to Asia but has adapted and grown very well in several regions of Brazil (Cruz, 1985). This species has important medicinal properties as it can be used to treat diabetes, reducing the urinary glycaemia. Despite of its medicinal importance, it has not been commercially cultivated since its propagation is negatively affected by the long time (10-12 month) taken between flowering and seed production (Lorenzi, 1992). The aspects mentioned above and the absence of ecological and genetic information give support to the development of rapid and efficient methods for Bauhinia multiplication through tissue culture. Organogenesis has been reported for some leguminous tree species such as Dalbergia sissoo (Kumar et al., 1991), Acacia nilotica subsp. indica (Dewan et al., 1992), Parkinsonia aculeata (Mathur & Mukunthakumar, 1992) and Sesbania grandifolia (Detrez et al., 1994). Although micropropagation and organogenesis have been reported for B. purpurea L. (Kumar, 1992), B. variegata (Mathur & Mukunthakumar, 1992) and B. vahlii Wight and Arnott (Upreti & Dhar, 1996),
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REVIEW Date palm micropropagation: Advances and applications Micropropagação de tamareira: Avanços e aplicações

REVIEW Date palm micropropagation: Advances and applications Micropropagação de tamareira: Avanços e aplicações

Light serves as an external factor to regulate the growth and development of in vitro plants. The broad spectrum of fluorescent lamps with a wavelength range from 380-750 nm are used as a light source for tissue culture (Kim et al., 2004). Light intensity and type of light affect date palm micropropagation (Al-Mayahi, 2016; Meziani et al., 2015). Meziani et al. (2015) evaluated the organogenesis of date palm cv. Mejhoul using different levels of light intensities. The results explained that the 2000-3000 lux light intensity enhances shoot elongation and greening but reduction in the shoot bud proliferation was observed. Darkness and low light intensity (500 lux) significantly induced advanced rooting. The 1000 lux light intensity during the multiplication stage showed optimal growth of culture with respect to shoot buds per explant, greening an advanced rooting. Recently Al-Mayahi (2016) conducted an experiment to test the effect of combinations of red + blue light emitting diode (18:2) (CRB-LED) and white fluorescent light on direct organogenesis by induction of adventitious buds from shoot tip and multiplication shoots of date palm cv. Alshakr. The results showed CRB-LED performed better in the production of shoot numbers than white fluorescent light. CRB-LED also significantly increased the total soluble carbohydrate, starch, free amino acids, peroxidase activity, potassium, magnesium and sodium contents of the in vitro shoots.
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Easy and efficient chemical sterilization of the culture medium for in vitro growth of gerbera using chlorine dioxide (ClO

Easy and efficient chemical sterilization of the culture medium for in vitro growth of gerbera using chlorine dioxide (ClO

Chemical sterilization represents nowadays an interesting alternative for culture medium sterilization, but in most of micropropagation laboratories autoclaving is considered the most security way for culture medium sterilization, main because the long repeatability of results and that avoid any type of microorganism resistance. However, the use of chemical sterilization with chlorine products, as chlorine dioxide, also represents an important tool to be use in plant tissue culture labs for production of thousands of plantlets, and represents one way to reduction costs of micropropagated plantlets, one of the challenges of micropropagation techniques for this century (CARDOSO et al. 2018). Studies with chemical sterilization of also liquid culture media for temporary immersion bioreactor systems needs be realized to evaluate the use and efficiency of chlorine dioxide also for this system, actually used for large-scale micropropagation of plantlets by tissue culture.
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Storage media for avulsed teeth: a literature review

Storage media for avulsed teeth: a literature review

literature showed that a wide array of types of wet storage media have been evaluated in laboratory studies and clinical reports, including cell and tissue culture solutions like Hank’s Balanced Salt Solution (HBSS); medical/hospital products developed specifically for organ storage purposes, such as Viaspan® and Euro-Collins®; culture media, like Minimum Essential Medium (MEM); saline; natural products like water, saliva, bovine milk and its variations, propolis, green tea, Morus rubra (red mulberry), egg white and coconut water; rehydrating solutions, like Gatorade® and Ricetral, and even contact lens solutions. Based on the literature, it could be stated that, so far, apart from Based on the literature, it could be stated that, so far, apart from solutions designed specifically for storage and culture purposes,, regular pasteurized whole milk is the most frequently recommended and with the best prognosis among other solutions that are likely to be available at the scene of an accident, such as water, saline or saliva. Its advantages include its high availability, ready accessibility, physiologically compatible pH and osmolality (fluid pressure) with the root-surface adhered PDL cells, presence of nutrients and growth factors. However, there is not yet a single solution that fulfills all requirements to be considered as the ideal medium for temporary storage of avulsed teeth, and research on this field should carry on.
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Amorphous calcium carbonate precipitation by cellular biomineralization in mantle cell cultures of Pinctada fucata.

Amorphous calcium carbonate precipitation by cellular biomineralization in mantle cell cultures of Pinctada fucata.

biogenic origin, and the expression and secretion of matrix proteins have also been detected in mantle explant cultures [21]. However, calcium carbonate polymorph deposited in mantle tissue culture and the cellular mechanisms of crystal formation remain unclear. Although haemocytes are thought to be directly involved in shell repair by storing intracellular calcium carbonate crystals and delivering crystals to the mineralization front [25], their contribution to normal shell formation is still under debate. Mantle cells are known to play central roles in shell and pearl formation. Considering the remarkable nacre structure and the contribution of the mantle cells to shell formation, the utilization of cell biological approaches is essential for further detailed analyses of shell formation mechanisms. Furthermore, these cells maintain cell-to-cell contacts in multicellular culture, may preserve the viability and functionality of mantle cells and may delay cellular aging and death, thus allowing in vitro biomineralization [22]. Hence, our focus is to identify the mechanism of aragonite nacreous layer formation by cellular biomineralization in vitro.
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Rev. bras. farmacogn.  vol.21 número2

Rev. bras. farmacogn. vol.21 número2

The main objective of the first publications on seaweed tissue culture was to investigate callus induction and growth in different species of red algae (Table 1). Polne-Fuller & Gibor (1987) described callus formation in nine species of rhodophytes, including species belonging to the genera Porphyra, Gelidium, Gracilaria, Eucheuma, Gigartina and Prionitis, but regeneration from callus was not observed. Similar results were reported by Gusev et al. (1987) for species of Ceramium, Laurencia, Gracilaria, Furcellaria and Phyllophora and also by Liu & Gordon (1987) for Pterocladiella capillacea. Other studies reported similar results for Porphyra umbilicalis (Liu &
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In vitro regeneration and antibacterial activity of Prunus domestica L.

In vitro regeneration and antibacterial activity of Prunus domestica L.

In tissue culture, PGRs are critical media components in determining the developmental pathway of plant. KIN (2 mg/l, 4 mg/l) obtained highest regeneration frequency (80%) as well as treatment without PGR was also gotten. Highest shoot length (1.20 ± 0.15 cm), highest shoot number (3.00±0.35) and highest leaf number (14.25±1.45) were observed at KIN 4.0 mg/l (Table 1). So, KIN is superior hormone among all hormones that were used. Highest mean number of shoots was observed 3.0 (Table 1) while it was observed 18.7 by Mante et al. (1989). Highest mean number of shoots were observed 1.87 (Table 1) recommended by Petri & Scorza (2009). Highest percentage of regenerating leafs was 14.25 (Table 1) while in another research paper it was observed 64.3 (Petri & Scorza, 2009).
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Respostas morfogenéticas de embriões de trigo em função do ambiente de cultivo da planta doadora de explantes

Respostas morfogenéticas de embriões de trigo em função do ambiente de cultivo da planta doadora de explantes

In 2005, there was no obvious clustering of climatic factors on the biplot (Figure 2c). In contrast to the pre- vious two years, rainfall had neither positive nor nega- tive effects on tissue culture traits. Also, minimum and maximum temperatures showed positive effects, while in previous two years their impacts on CF, RC and PPE were mainly negative. The reason for this may be be- cause 2005 was the coolest year, particularly in the pe- riod from flowering to the medium milk stage (Table 2). It may be possible to increase competence of imma- ture wheat embryos for plant regeneration by growing donor plants under cool conditions, which appears to delay the accumulation of endogenous hormone levels in kernels (Hess and Carman, 1998). The callus forma- tion was positively influenced by varT10, minT6, maxT10, minT10, as well as SH10. The regeneration ca- pacity also had a positive correlation with the tempera- ture based factors (varT6, maxT2, minT2, varT2) and the sunshine duration (SH2 and SH6). For barley, a sufficient regeneration from greenhouse-grown donor plants was obtained when temperatures were moderate and natu- ral light levels were high (Dahleen, 1999). Factors posi- tively correlated to CF and regenerative potential were mainly effective during ten and two weeks prior to sam- pling of embryos, respectively.
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