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Adventive in vitro regenerants of Populus x berolinensis Dipp. appeared after transfer of conglomerates on elongation (P2) medium. The regenerant with greatest height is seen. Diameter of jar-6 cm.
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Adventive regenerants of Populus × berolinensis Dipp. were obtained on 1/2 MS salts with sucrose, vitamins, BA, TDZ and NAA using stem, petiole and leaf explants. They differed from each other in plantlet dimensions. More than 50 clones were produced from initial regenerants by excision and rooting of apexes and lateral shoots. Differences in stem...
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... Although, somaclonal variation in trees is usually undesirable (Tremblay et al., 1999;Breton et al. 2006;Marum et al., 2009), a few useful somaclonal variants have been reported in tree species (Ahuja, 1998;Bairu et al., 2011;Krishna et al., 2016). Some of these useful variants include: height and morphological traits in poplar (Fry et al., 1997) and olive (Leva et al., 2012); disease resistance in poplar (Ostry and Ward, 2003), apple citrus, and mango (Krishna et al., 2016); different growth rates in poplar (Gamburg and Voinikov, 2013); and stress tolerance in radiata pine (Montalbán et al., 2014). ...
Woody plants have been cultured in vitro since the 1930s. After that time much progress has been made in the culture of tissues, organs, cells, and protoplasts in tree species. Tree biotechnology has been making strides in clonal propagation by organogenesis and somatic embryogenesis. These regeneration studies have paved the way for gene transfer in forest trees. Transgenics from a number of forest tree species carrying a variety of recombinant genes that code for herbicide tolerance, pest resistance, lignin modification, increased woody bio-mass, and flowering control have been produced by Agrobacterium -mediated and biolistic methods, and some of them are undergoing confined field trials. Although relatively stable transgenic clones have been produced by genetic transformation in trees using organogenesis or somatic embryogenesis, there were also unintended unstable genetic events. In order to overcome the problems of randomness of transgene integration and instability reported in Agrobacterium -mediated or biolistically transformed plants, site-specific transgene insertion strategies involving clustered regularly interspaced short palindromic repeats (CRISPR-Cas9) platform offer prospects for precise genome editing in plants. Nevertheless, it is important to monitor phenotypic and genetic stability of clonal material, not just under greenhouse conditions, but also under natural field conditions. Genetically modified poplars have been commercialized in China, and eucalypts and loblolly pine are expected to be released for commercial deployment in USA. Clonal forestry and transgenic forestry have to cope with rapid global climate changes in the future. Climate change is impacting species distributions and is a significant threat to biodiversity. Therefore, it is important to deploy Strategies that will assist the survival and evolution of forest tree species facing rapid climate change. Assisted migration (managed relocation) and biotechnological approaches offer prospects for adaptation of forest trees to climate change.
... The occurrence of somaclonal variation of in vitro-grown plantlets is a common problem (Gamburg and Voinikov 2013;Krishna et al. 2016) making it necessary to ensure the regenerants are true-to-type. To assure the genetic fidelity of regenerated plantlets several molecular markers have been used, such as RAPD (Bublyk et al. 2012), ISSR (Bramhanapalli et al. 2017), SSR & AFLP (Tiwari et al. 2013). ...
Solanum viarum Dunal is an important medicinal plant with a high quantity of steroidal alkaloids used for the synthesis of contraceptives, corticosteroids, and sex hormones. It is also used by Indian tribal people for the treatment of leprosy, toothache, and diabetes. Therefore, to meet the existing needs for this plant, it is necessary to develop an efficient regeneration system useful for rapid and large-scale clonal propagation with ensured genetic fidelity. An efficient and improved regeneration protocol for prickly and prickleless genotypes of S. viarum has been developed using three explants, leaf, petiole, and internodes, under the influence of two plant growth regulators, thidiazuron (TDZ) and 6-benzyladenine (BA). Effects of genotype, explant type, and concentrations of TDZ and BA were studied. A higher percentage of shoot organogenesis (78.25% ± 2.53) and shoot number per explant (6.79 ± 1.04) were achieved in the leaf segments of prickly genotype cultured on modified Murashige and Skoog (MS) medium supplemented with TDZ (1.50 mg L−1). Furthermore, basal leaf segments showed 100% regeneration from the prickly genotype. A significantly higher content of total phenolics was quantified in prickleless (3.66 μg mg−1) than prickly genotypes (2.73 μg mg−1). The monomorphic banding pattern of random amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR) analysis confirmed the genetic fidelity of the regenerated plants. Additionally, flow cytometric analysis of regenerants showed no variation in the ploidy levels when compared to the mother (control) plants. These results clearly depicted the efficiency of developed protocol that can be utilized for generating genetically stable population of S. viarum.
... We also observed significant differences in transformation efficiency when using clone 717 in vitro cultures that differed in age. The low efficiency obtained with the 7-year-old cultures may result from tissue culture-specific somaclonal variation (Larkin and Scowcroft 1981), which happens at a low rate in Populus ) but has been reported in several hybrid clones and can lead to changes in leaf morphology and growth rate (Son et al. 1993;Gamburg and Voinikov 2013;Thakur and Ishii 2014). It is also possible that naturally occurring variations, similar to what we observed in PtAG1, led to different transformation competence between the two sets of clone 717 cultures. ...
Gene flow from recombinant-DNA-modified (GMO) trees is a major barrier to their public acceptance and regulatory approval. Because many intensively grown trees are vegetatively propagated, complete sexual sterility could be a powerful means to mitigate or prevent gene flow. We tested four pairs of zinc-finger nucleases (ZFNs) as mutagenic agents against the LEAFY and AGAMOUS orthologs in poplar that are expected to be required for sexual fertility. To reduce the potential for pleiotropic effects from mutagenesis, each of the pairs was functionally linked to a heat shock promoter to provide inducible ZFN expression. Using Agrobacterium tumefaciens, we transformed more than 21,000 total explants compromised of both male and female hybrid poplar. The rate of transformation for the ZFN constructs (2 %) was generally reduced compared to the transgenic control (8 %). We produced 391 ZFN transgenic shoots of which only two developed into plants with mutations in a target gene; both were 7-bp deletions in one allele of the PtAG2 locus. No mutations were observed in the PtAG1 or PtLFY loci. Our results indicate a mutation rate of zero to 0.3 % per explant per allele, among the lowest reported for ZFN mutagenesis in plants. The combined effects of low recovery of transgenic plants, a modest mutation frequency, and much higher reported rates of directed mutation for other gene editing methods suggest that the efficient use of ZFNs in poplar requires further technical improvements.
... Recent applications of multivariate analysis on morphometric and meristic data in the study of morphological variation in plants include Valenzuela et al. (2011) andPlazas et al. (2014). Similar approaches have been used for other tissue cultured plants such as tomato (Pratta et al., 2000), soybean (Radhakrishnan and Kumari, 2008) banana (Sheidai et al., 2008) and poplar tree (Gamburg and Voinikov, 2013). ...
... The use of cluster analysis to complement either principal coordinate or principal component analyses of morphological or genetic traits in plants is well documented (Pratta et al., 2000;Song et al., 2000;Sheidai et al., 2008Sheidai et al., , 2010Valenzuela et al., 2011;Abdellatif et al., 2012;Gamburg and Voinikov, 2013;Plazas et al., 2014). In the present study cluster analysis validated the PCOORDA plots. ...
Somaclonal variation induced in vitro during tissue culture can be a problem in clonal micropropagation of elite plants. This study investigated the extent of morphological and genetic similarity or dissimilarity between Melia volkensii in vitro plants (somatic seedlings) obtained via somatic embryogenesis and normal seedlings. Comparisons were made between in vitro plants regenerated directly from cotyledon explants, indirectly from zygotic embryos and normal seedlings of the same parent trees. Regeneration was achieved using half MS medium supplemented with 0.05 mg/l thidiazuron. Shoots were elongated in half MS with 0.1 mg/l BAP plus 0.01 mg/l IAA then rooted in half MS with 0.1 mg/l IBA and 0.1 mg/l NAA. Six morphometric and five meristic characters were used for the morphological characterization. PCR-RAPD markers were used for assessment of genetic similarity or distance. Multivariate analysis using principal coordinates, cluster analysis, analysis of similarities (Anosim) and similarity percentages analysis (SIMPER) revealed significant dissimilarities (p< 0.0001) in morphometric and meristic characters between the in vitro plants and normal seedlings. However, significant similarity (p<0.01) was observed in the RAPD-genic characters of the in vitro plants and normal seedlings. Out of six morphometric traits, taproot length, internode length and shoot height were the most important sources of dissimilarity, cumulatively accounting for 72.37% of overall morphometric dissimilarity. Number of lateral roots was the single most important source of meristic dissimilarity, with 77.02% contribution. Plants regenerated directly from cotyledons were more similar to the normal seedlings in morphological and RAPD-marker characters than those regenerated indirectly from zygotic embryos. This study paves the way for identification of trait-specific RAPD markers for further characterization through sequence-characterized amplified regions (SCARs).
... Recent applications of multivariate analysis on morphometric and meristic data in the study of morphological variation in plants include Valenzuela et al. (2011) andPlazas et al. (2014). Similar approaches have been used for other tissue cultured plants such as tomato (Pratta et al., 2000), soybean (Radhakrishnan and Kumari, 2008) banana (Sheidai et al., 2008) and poplar tree (Gamburg and Voinikov, 2013). ...
... Zygotic embryo clones had 36.4% genetic differentiation from wild type whilst the value for cotyledon clones was 28.4%, making cotyledon clones to be the closest in genetic similarity to the wild type seedlings. The use of cluster analysis to complement either principal coordinate or principal component analyses of morphological or genetic traits in plants is well documented (Pratta et al., 2000;Song et al., 2000;Sheidai et al., 2008Sheidai et al., , 2010Valenzuela et al., 2011;Abdellatif et al., 2012;Gamburg and Voinikov, 2013;Plazas et al., 2014). In the present study cluster analysis validated the PCOORDA plots. ...
Somaclonal variation induced in vitro during tissue culture can be a problem in clonal micropropagation of elite plants. This study investigated the extent of morphological and genetic similarity or dissimilarity between Melia volkensii in vitro plants (somatic seedlings) obtained via somatic embryogenesis and normal seedlings. Comparisons were made between in vitro plants regenerated directly from cotyledon explants, indirectly from zygotic embryos and normal seedlings of the same parent trees. Regeneration was achieved using half MS medium supplemented with 0.05 mg/l thidiazuron. Shoots were elongated in half MS with 0.1 mg/l BAP plus 0.01 mg/l IAA then rooted in half MS with 0.1 mg/l IBA and 0.1 mg/l NAA. Six morphometric and five meristic characters were used for the morphological characterization. PCR-RAPD markers were used for assessment of genetic similarity or distance. Multivariate analysis using principal coordinates, cluster analysis, analysis of similarities (Anosim) and similarity percentages analysis (SIMPER) revealed significant dissimilarities (p< 0.0001) in morphometric and meristic characters between the in vitro plants and normal seedlings. However, significant similarity (p<0.01) was observed in the RAPD-genic characters of the in vitro plants and normal seedlings. Out of six morphometric traits, taproot length, internode length and shoot height were the most important sources of dissimilarity, cumulatively accounting for 72.37% of overall morphometric dissimilarity. Number of lateral roots was the single most important source of meristic dissimilarity, with 77.02% contribution. Plants regenerated directly from cotyledons were more similar to the normal seedlings in morphological and RAPD-marker characters than those regenerated indirectly from zygotic embryos. This study paves the way for identification of trait-specific RAPD markers for further characterization through sequence-characterized amplified regions (SCARs).
