Somatic embryogenesis and plantlet regeneration from mature zygotic embryos of Manchurian ash (Fraxinus mandshurica Rupr.)

Yang, Ling; Bian, Lei; Shen, Hailong; Li, Yuhua

doi:10.1007/s11240-013-0345-8

Cited by 20 publications

(14 citation statements)

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“…8). This nding is different from those obtained from the regeneration of other F. mandshurica [13] or ashes [27], indicating that the regeneration rate of F. mandshurica hypocotyls not only depends on the starting hypocotyl explants and time of pretreatment, but also depends on the supplement used in pretreatment and other external stimulations.…”

Section: Discussioncontrasting

confidence: 70%

“…During the grafting process, the degree of joining between scions and rootstocks, the quality of scions, and the method of grafting all affect the nal survival rate of F. mandshurica [9]. Studies on tissue culture of F. mandshurica, such as callus culture [10], somatic embryogenesis [11][12][13], and axillary bud proliferation [14], all demonstrated certain limitations including high rates of adventitious buds browning and somatic embryo deformity, and low rate of shoot elongation. A variety of materials including the vegetative organs and reproductive organs of Fraxinus mandshurica are cultivated, but the best results can only obtain the growth of a single bud, and have not obtained any hyperplastic plants [12].Genetic engineering technologies provide an attractive way to effectively improve current varieties of F. mandshurica [15].…”

Section: Introductionmentioning

confidence: 99%

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A Rapid and Efficient Protocol for Adventitious Shoot Regeneration and Genetic Transformation of Manchurian Ash (Fraxinus mandshurica Rupr.) using Hypocotyl Explants

Li¹,

Cao²,

Yu³

et al. 2020

Preprint

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Background: Manchurian ash (Fraxinus mandshurica Rupr.) is an endangered hardwood tree species, providing both economic and medicinal benefits. However, observations such as browning of adventitious shoot buds and high rate of somatic embryo abnormality, were presented in protocols of F. mandshurica regeneration. Therefore, a rapid and high-efficiency regeneration system is demanded for mass propagation and genetic transformation of F. mandshurica.Results: We have developed an efficient regeneration system through adventitious shoot organogenesis in F. mandshurica using hypocotyl explants, with which the adventitious shoots are able to elongate and were obtained in an affordable time. Hypocotyls excised from embryos were pre-cultured in the dark on woody plant medium (WPM) supplemented with 6 g L -1 potassium citrate, and then inoculated on WPM medium supplemented with different concentrations of plant growth regulators (PGRs) to induce adventitious shoot bud formation. The induction medium supplemented with a single PGR of 1.0 mg L -1 thidiazuron (TDZ) was the best treatment, showing 86.67% shoot bud induction with a 15-day initial dark culture, followed by culture under a low light condition. The survival rate of regenerated shoot buds reached 70.97% on WPM medium supplemented with 0.025 mg L -1 TDZ and 1.0 mg L -1 gibberellic acid (GA3). Based on this regeneration system, By using the sonication plus vacuum-infiltration method,a protocol for Agrobacterium tumefaciens -mediated transformation of hypocotyls was established,the transformation rate was determined to be 3.57%. Conclusions: Key factors, such as the potassium citrate pretreatment, wound treatment on explants, variable light conditions, and significant PGR interactions, were revealed to affect the induction and elongation of adventitious shoots from F. mandshurica hypocotyls in this study. The adventitious shoots, tissue culture plantlets, and rooted plantlets were obtained at 40, 80-100, and 160 days, respectively. This regeneration system shortens the period of traditional regeneration methods, which require months to induce callus from leaves or stems, and additional several months for organ differentiation. In addition, the Agrobacterium-mediated transformation protocol established on the basis of this regeneration system provides a foundation for breeding, genetic improvement and genomic studies of F. mandshurica.

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Section: Discussioncontrasting

confidence: 70%

Section: Introductionmentioning

confidence: 99%

A Rapid and Efficient Protocol for Adventitious Shoot Regeneration and Genetic Transformation of Manchurian Ash (Fraxinus mandshurica Rupr.) using Hypocotyl Explants

Li¹,

Cao²,

Yu³

et al. 2020

Preprint

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“…Asexual reproduction of individuals who have been selected, improved, and genetically manipulated can accelerate the breeding process (Lelu-Walter et al 2013). Over the last 17 years, studies on the somatic embryogenesis of F. mandshurica have identified good explant sources for somatic embryogenesis and the optimum period for explant harvesting (Sun et al 2010), described the physiological and biochemical changes in somatic embryogenesis (Cong et al 2012), SE maturation, and germination (Yang et al 2013), documented the proteomic profile of SEs (Liu et al 2015). In our research, we have found that SEs of F. mandshurica can form directly or indirectly via callus.…”

Section: Introductionmentioning

confidence: 99%

“…The main problems with direct somatic embryogenesis of F. mandshurica are the low SE yield (Yang et al 2013), unsynchronized SE development (Zhang and Shen 2007;Yang et al 2013), and genetic instability, all of which restrict large-scale production. In this study, we devised a strategy to increase callus proliferation, improve embryo differentiation, and synchronize embryo development, which ultimately increased the number of F. mandshurica SEs and complete regenerated plants.…”

Section: Introductionmentioning

confidence: 99%

Indirect somatic embryogenesis and regeneration of Fraxinus mandshurica plants via callus tissue

et al. 2020

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supplemented with 0.15 mg•L −1 naphthalene acetic acid. The highest callus proliferation coefficient (240.5) was obtained on McCown's Woody Plant Medium containing 0.1 mg•L −1 6-benzyl adenine and 0.15 mg•L −1 2, 4-dichlorophenoxyacetic acid. The highest number of SEs (1020.5 g −1 fresh weight) was obtained on MS½ medium supplemented with 1 mg•L −1 6-benzyladenine. The highest number of cotyledon embryos (397/g fresh weight) was obtained by incubating materials on medium containing 1 mg•L −1 abscisic acid and then applying a drying treatment. The cotyledon embryos were milky white, uniformly sized (average length 4.7 mm), and 80% of them were normal. The SE rooting percentage on ½MS medium containing 0.01 mg•L −1 NAA was 37.5%. Overall, the germination percentage of SEs was 26.4%, and complete regenerated plants were obtained after transplanting and acclimation. These results provide more possibilities for the preservation and breeding of F. mandshurica.

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“…However, the regeneration efficiency of a great number of plant species is still low under in vitro conditions. Embryogenic callus formation offers great potential for fostering in vitro culture efficiency whereby somatic cells are induced to differentiate embryogenic cells and form somatic embryos that develop into new plants [ 2 , 3 ]. The ability to initiate embryogenic cultures is controlled by the intrinsic and external factors of the explants, including the genotype, the developmental stage, and the different tissue of the explants, as well as the factors associated with the medium as perceived by the complementary sensors in cells, including stress, phytohormones, and the artificial nutritional environment in the culture process [ 4 – 6 ].…”

Section: Introductionmentioning

confidence: 99%

De novo assembly and comparative analysis of the transcriptome of embryogenic callus formation in bread wheat (Triticum aestivum L.)

et al. 2017

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BackgroundDuring asexual reproduction the embryogenic callus can differentiate into a new plantlet, offering great potential for fostering in vitro culture efficiency in plants. The immature embryos (IMEs) of wheat (Triticum aestivum L.) are more easily able to generate embryogenic callus than mature embryos (MEs). To understand the molecular process of embryogenic callus formation in wheat, de novo transcriptome sequencing was used to generate transcriptome sequences from calli derived from IMEs and MEs after 3d, 6d, or 15d of culture (DC).ResultsIn total, 155 million high quality paired-end reads were obtained from the 6 cDNA libraries. Our de novo assembly generated 142,221 unigenes, of which 59,976 (42.17%) were annotated with a significant Blastx against nr, Pfam, Swissprot, KOG, KEGG, GO and COG/KOG databases. Comparative transcriptome analysis indicated that a total of 5194 differentially expressed genes (DEGs) were identified in the comparisons of IME vs. ME at the three stages, including 3181, 2085 and 1468 DEGs at 3, 6 and 15 DC, respectively. Of them, 283 overlapped in all the three comparisons. Furthermore, 4731 DEGs were identified in the comparisons between stages in IMEs and MEs. Functional analysis revealed that 271transcription factor (TF) genes (10 overlapped in all 3 comparisons of IME vs. ME) and 346 somatic embryogenesis related genes (SSEGs; 35 overlapped in all 3 comparisons of IME vs. ME) were differentially expressed in at least one comparison of IME vs. ME. In addition, of the 283 overlapped DEGs in the 3 comparisons of IME vs. ME, excluding the SSEGs and TFs, 39 possessed a higher rate of involvement in biological processes relating to response to stimuli, in multi-organism processes, reproductive processes and reproduction. Furthermore, 7 were simultaneously differentially expressed in the 2 comparisons between the stages in IMEs, but not MEs, suggesting that they may be related to embryogenic callus formation. The expression levels of genes, which were validated by qRT-PCR, showed a high correlation with the RNA-seq value.ConclusionsThis study provides new insights into the role of the transcriptome in embryogenic callus formation in wheat, and will serve as a valuable resource for further studies addressing embryogenic callus formation in plants.Electronic supplementary materialThe online version of this article (10.1186/s12870-017-1204-2) contains supplementary material, which is available to authorized users.

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Somatic embryogenesis and plantlet regeneration from mature zygotic embryos of Manchurian ash (Fraxinus mandshurica Rupr.)

Cited by 20 publications

References 31 publications

A Rapid and Efficient Protocol for Adventitious Shoot Regeneration and Genetic Transformation of Manchurian Ash (Fraxinus mandshurica Rupr.) using Hypocotyl Explants

A Rapid and Efficient Protocol for Adventitious Shoot Regeneration and Genetic Transformation of Manchurian Ash (Fraxinus mandshurica Rupr.) using Hypocotyl Explants

Indirect somatic embryogenesis and regeneration of Fraxinus mandshurica plants via callus tissue

De novo assembly and comparative analysis of the transcriptome of embryogenic callus formation in bread wheat (Triticum aestivum L.)

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