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11 pages, 5595 KiB

Open AccessArticle

Spontaneous Regeneration of Plantlets Derived from Hairy Root Cultures of Lopezia racemosa and the Cytotoxic Activity of Their Organic Extracts

by Norely Vargas-Morales, Norma Elizabeth Moreno-Anzúrez, Janeth Téllez-Román, Irene Perea-Arango, Susana Valencia-Díaz, Alfonso Leija-Salas, Edgar R. Díaz-García, Pilar Nicasio-Torres, María Del Carmen Gutiérrez-Villafuerte, Jaime Tortoriello-García and Jesús Arellano-García

Plants 2022, 11(2), 150; https://doi.org/10.3390/plants11020150 - 06 Jan 2022

Cited by 6 | Viewed by 1696

Abstract

A histological analysis was performed with the aim of elucidating the spontaneous regeneration process of the hairy root lines LRT 2.3 and LRT 6.4, derived from Lopezia racemosa leaf explants and genetically transformed with the Agrobacterium rhizogenes strain ATCC15834/pTDT. The analysis showed [...] Read more.

A histological analysis was performed with the aim of elucidating the spontaneous regeneration process of the hairy root lines LRT 2.3 and LRT 6.4, derived from Lopezia racemosa leaf explants and genetically transformed with the Agrobacterium rhizogenes strain ATCC15834/pTDT. The analysis showed both lines regenerate via indirect somatic embryogenesis; LRT 6.4 also regenerated by direct organogenesis. The morphogenic characteristics of the regenerated plantlets from both lines showed the typical characteristics, described previously, including a higher number of axillary shoot formation, short internodes, and plagiotropic roots compared with wild-type seedlings. The regeneration process occurred without the addition of plant growth regulators and was linked to the sucrose concentration in the culture medium. Reducing the sucrose concentration from 3% to 2%, 1%, and 0.5% increased the regeneration rate in LRT 6.4; the effect was less pronounced in LRT 2.3. The cytotoxic activity of different organic extracts obtained from roots and shoots were evaluated in the cancer cell lines HeLa (cervical carcinoma), HCT-15 (colon adenocarcinoma), and OVCAR (ovary carcinoma). The hexane and dichloromethane extracts from roots of both lines showed cytotoxic activity against the HeLa cell line. Only the dichloromethane extract from the roots of PLRT 2.3 showed cytotoxic activity against the OVCAR cell line. None of the methanol extracts showed cytotoxic activity, nor the shoot extracts from any solvent. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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13 pages, 1324 KiB

Open AccessArticle

In Vitro Regeneration of Miscanthus x giganteus through Indirect Organogenesis: Effect of Explant Type and Growth Regulators

by Aušra Blinstrubienė, Inga Jančauskienė and Natalija Burbulis

Plants 2021, 10(12), 2799; https://doi.org/10.3390/plants10122799 - 17 Dec 2021

Cited by 2 | Viewed by 1998

Abstract

Miscanthus x giganteus is a spontaneous sterile hybrid therefore the creation of useful genetic diversity by conventional breeding methods is restricted. Plant regeneration through indirect organogenesis may be a useful approach to create genetic variability of this important agricultural crop. The present study [...] Read more.

Miscanthus x giganteus is a spontaneous sterile hybrid therefore the creation of useful genetic diversity by conventional breeding methods is restricted. Plant regeneration through indirect organogenesis may be a useful approach to create genetic variability of this important agricultural crop. The present study aimed to evaluate the effect of the explant type and growth regulators on indirect organogenesis of Miscanthus x giganteus and to determine the ploidy level of plant regenerants by flow cytometry. On average, the highest percentage of morphogenic callus tested explants formed in the medium supplemented with 2.5 mg L^–1 IBA + 0.1 mg L^–1 BAP + 4.0 mg L^–1 l-proline. The most intensive secondary differentiation of callus cells was observed in the medium supplemented with 4.0 mg L^–1 ZEA + 1.0 mg L^–1 NAA. The highest root formation frequency with the highest number of roots was determined in the MS nutrient medium supplemented with 0.4 mg L^–1 IBA, where more than 95% of plant regenerants survived and were growing normally. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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14 pages, 7314 KiB

Open AccessArticle

Establishment of a Novel and Efficient Agrobacterium-Mediated in Planta Transformation System for Passion Fruit (Passiflora edulis)

by Hafiz Muhammad Rizwan, Qiang Yang, Ahmed Fathy Yousef, Xiaoxue Zhang, Yasir Sharif, Jia Kaijie, Meng Shi, Han Li, Nigarish Munir, Xuelian Yang, Xiaoxia Wei, Ralf Oelmüller, Chunzhen Cheng and Faxing Chen

Plants 2021, 10(11), 2459; https://doi.org/10.3390/plants10112459 - 15 Nov 2021

Cited by 7 | Viewed by 3039

Abstract

Passion fruit (Passiflora edulis) is an important fruit crop with high economic value. Genetic engineering plays an important role in crop improvement with desired traits and gene functional studies. The lack of a simple, efficient, and stable transformation system for passion [...] Read more.

Passion fruit (Passiflora edulis) is an important fruit crop with high economic value. Genetic engineering plays an important role in crop improvement with desired traits and gene functional studies. The lack of a simple, efficient, and stable transformation system for passion fruit has greatly limited gene functional studies. In this study, a simple and efficient Agrobacterium-mediated in planta transformation system for passion fruit was established, using Agrobacterium virulent strain EHA105 harboring the binary vectors pCAMBIA1301 and pCAMBIA1302 with GUS and GFP reporter genes. The system requires less time and labor costs than conventional transformation systems, and no additional phytohormones and sterile conditions are required. Regeneration efficiency of 86% and transformation efficiency of 29% were achieved, when the wounds were wrapped with Parafilm and the plants were kept in darkness for 15 days. Approximately 75% of the regenerated plants had a single shoot and 26% multiple shoots. The transformation was confirmed at the DNA and RNA levels as well as by GUS staining and GFP fluorescent measurements. The developed protocol will contribute to the genetic improvement of passion fruit breeding. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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9 pages, 1714 KiB

Open AccessArticle

Temporal Control of Morphogenic Factor Expression Determines Efficacy in Enhancing Regeneration

by Juan H. Gonzalez, Joseph S. Taylor, Kelsey M. Reed, R. Clay Wright and Bastiaan O. R. Bargmann

Plants 2021, 10(11), 2271; https://doi.org/10.3390/plants10112271 - 23 Oct 2021

Cited by 4 | Viewed by 2361

Abstract

Background: Regeneration of fertile plants from tissue culture is a critical bottleneck in the application of new plant breeding technologies. Ectopic overexpression of morphogenic factors is a promising workaround for this hurdle. Methods: Conditional overexpression of WUS and ARF5Δ was used to [...] Read more.

Background: Regeneration of fertile plants from tissue culture is a critical bottleneck in the application of new plant breeding technologies. Ectopic overexpression of morphogenic factors is a promising workaround for this hurdle. Methods: Conditional overexpression of WUS and ARF5Δ was used to study the effect of timing the overexpression of these morphogenic factors during shoot regeneration from root explants in Arabidopsis. In addition, their effect on auxin-signaling activation was examined by visualization and cytometric quantification of the DR5:GFP auxin-signaling reporter in roots and protoplasts, respectively. Results: The induced expression of both WUS and ARF5Δ led to an activation of auxin signaling in roots. Activation of auxin signaling by WUS and ARF5Δ was further quantified by transient transformation of protoplasts. Ectopic overexpression of both WUS and ARF5Δ enhanced regeneration efficiency, but only during the shoot-induction stage of regeneration and not during the callus-induction stage. Conclusions: The overexpression of WUS and ARF5Δ both lead to activation of auxin signaling. Expression during the shoot-induction stage is critical for the enhancement of shoot regeneration by WUS and ARF5Δ. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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12 pages, 2120 KiB

Open AccessArticle

Application of Ascophyllum nodosum-Based Soluble Extract on Micropropagation and Regeneration of Nicotiana benthamiana and Prunus domestica

by Mohamed Faize, Lydia Faize, Lorenzo Burgos, Alan T. Critchley and Nuria Alburquerque

Plants 2021, 10(7), 1354; https://doi.org/10.3390/plants10071354 - 02 Jul 2021

Cited by 5 | Viewed by 2649

Abstract

In the present study, the effect of a commercial extract of the seaweed Ascophyllum nodosum on in vitro micropropagation, shoot regeneration, and rhizoghenesis were studied in Nicotiana benthamiana and Prunus domestica. Results showed that the MS medium supplemented with various concentrations of [...] Read more.

In the present study, the effect of a commercial extract of the seaweed Ascophyllum nodosum on in vitro micropropagation, shoot regeneration, and rhizoghenesis were studied in Nicotiana benthamiana and Prunus domestica. Results showed that the MS medium supplemented with various concentrations of the Ascophyllum extract (5, 10, 50, and 100 mg L⁻¹) significantly enhanced the number of regenerated buds from N. benthamiana leaf discs to the conventional MS regenerating medium. Increases ranged from 3.5 to 6.5 times higher than the control. The effect of the Ascophyllum extract on N. benthamiana micropropagation was assessed through the measurement of some plant growth parameters. Results showed that the extract alone could not replace the micropropagation medium since shoot length, shoot diameter, root length, and leaf area were significantly reduced. However, its combination with a half-strength MS medium enhanced these parameters. Its effect was also evaluated on regeneration from plum hypocotyl slices. When added to the shoot regeneration medium without any plant growth regulators, the Ascophyllum extract alone could induce shoot regeneration. However, the percentage of bud regeneration and number of regenerated buds were lower than with the conventional shoot regeneration medium containing complete growth regulators. In contrast, the Ascophyllum extract drastically promoted rhizogenesis from plum hypocotyl slices. These results pave the way for the possible use of A. nodosum extracts in in vitro mass propagation of higher plants. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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13 pages, 3638 KiB

Open AccessArticle

Induction of Polyploidy and Metabolic Profiling in the Medicinal Herb Wedelia chinensis

by Yung-Ting Tsai, Po-Yen Chen and Kin-Ying To

Plants 2021, 10(6), 1232; https://doi.org/10.3390/plants10061232 - 17 Jun 2021

Cited by 15 | Viewed by 3182

Abstract

Wedelia chinensis, which belongs to the Asteraceae family, is a procumbent, perennial herb. It has medicinal anti-inflammatory properties and has been traditionally used as folk medicine in East and South Asia for treating fever, cough and phlegm. In Taiwan, W. chinensis is [...] Read more.

Wedelia chinensis, which belongs to the Asteraceae family, is a procumbent, perennial herb. It has medicinal anti-inflammatory properties and has been traditionally used as folk medicine in East and South Asia for treating fever, cough and phlegm. In Taiwan, W. chinensis is a common ingredient of herbal tea. Previous studies showed that the plant leaves contain four major bioactive compounds, wedelolactone, demethylwedelolactone, luteolin and apigenin, that have potent antihepatoxic activity, and are thus used as major ingredients in phytopharmaceutical formulations. In this study, we set up optimal conditions for induction of ploidy in W. chinensis. Ploidy can be an effective method of increasing plant biomass and improving medicinal and ornamental characteristics. By using flow cytometry and chicken erythrocyte nuclei as a reference, the DNA content (2C) or genome size of W. chinensis was determined to be 4.80 picograms (pg) in this study for the first time. Subsequently, we developed the successful induction of five triploid and three tetraploid plants by using shoot explants treated with different concentrations (0, 0.25, 0.5, 1, 1.5, 2 g/L) of colchicine. No apparent morphological changes were observed between these polyploid plants and the diploid wild-type (WT) plant, except that larger stomata in leaves were found in all polyploid plants as compared to diploid WT. Ultra-performance liquid chromatography coupled with tandem mass spectrometry was used to quantify the four index compounds (wedelolactone, demethylwedelolactone, luteolin, apigenin) in these polyploid plants, and fluctuating patterns were detected. This is the first report regarding polyploidy in the herbal plant W. chinensis. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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12 pages, 1915 KiB

Open AccessArticle

Citrus Cell Suspension Culture Establishment, Maintenance, Efficient Transformation and Regeneration to Complete Transgenic Plant

by M. Moniruzzaman, Yun Zhong, Zhifeng Huang, Huaxue Yan, Lv Yuanda, Bo Jiang and Guangyan Zhong

Plants 2021, 10(4), 664; https://doi.org/10.3390/plants10040664 - 30 Mar 2021

Cited by 13 | Viewed by 4672

Abstract

Agrobacterium-mediated transformation of epicotyl segment has been used in Citrus transgenic studies. The approach suffers, however, from limitations such as occasionally seed unavailability, the low transformation efficiency of juvenile tissues and the high frequency of chimeric plants. Therefore, a suspension cell culture [...] Read more.

Agrobacterium-mediated transformation of epicotyl segment has been used in Citrus transgenic studies. The approach suffers, however, from limitations such as occasionally seed unavailability, the low transformation efficiency of juvenile tissues and the high frequency of chimeric plants. Therefore, a suspension cell culture system was established and used to generate transgenic plants in this study to overcome the shortcomings. The embryonic calli were successfully developed from undeveloped ovules of the three cultivars used in this study, “Sweet orange”-Egyptian cultivar (Citrus sinensis), “Shatangju” (Citrus reticulata) and “W. Murcott” (Citrus reticulata), on three different solid media. Effects of media, genotypes and ages of ovules on the induction of embryonic calli were also investigated. The result showed that the ovules’ age interferes with the callus production more significantly than media and genotypes. The 8 to 10 week-old ovules were found to be the best materials. A cell suspension culture system was established in an H+H liquid medium. Transgenic plants were obtained from Agrobacterium-mediated transformation of cell suspension as long as eight weeks subculture intervals. A high transformation rate (~35%) was achieved by using our systems, confirming BASTA selection and later on by PCR confirmation. The results demonstrated that transformation of cell suspension should be more useful for the generation of non-chimeric transgenic Citrus plants. It was also shown that our cell suspension culture procedure was efficient in maintaining the vigor and regeneration potential of the cells. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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Review

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21 pages, 847 KiB

Open AccessEditor’s ChoiceReview

Polyploidization in Orchids: From Cellular Changes to Breeding Applications

by Joe Abdul Vilcherrez-Atoche, Carla Midori Iiyama and Jean Carlos Cardoso

Plants 2022, 11(4), 469; https://doi.org/10.3390/plants11040469 - 09 Feb 2022

Cited by 14 | Viewed by 4158

Abstract

Polyploidy occurs naturally in plants through cell division errors or can artificially be induced by antimitotic agents and has ecological effects on species adaptation, evolution, and development. In agriculture, polyploidy provides economically improved cultivars. Furthermore, the artificial induction of polyploids increases the frequency; [...] Read more.

Polyploidy occurs naturally in plants through cell division errors or can artificially be induced by antimitotic agents and has ecological effects on species adaptation, evolution, and development. In agriculture, polyploidy provides economically improved cultivars. Furthermore, the artificial induction of polyploids increases the frequency; thus, it accelerates obtaining polyploid plants used in breeding programs. This is the reason for its use in developing many crops of economic interest, as is the case of orchids in the flower market. Polyploidy in ornamental plants is mainly associated with flowers of larger size, fragrance, and more intense coloring when compared to naturally diploid plants. Currently, orchids represent the largest flower market worldwide; thus, breeding programs aim to obtain flowers with the larger size, durability, intense colors, and resistance to pathogens. Furthermore, orchid hybridization with polyploidy induction has been used to produce improved hybrid cultivars. Thus, the objective of this review was to compile information regarding the natural occurrence, importance, and methods of induction of polyploidy in orchids. The study also summarizes the significance of polyploids and techniques associated with artificially inducing polyploidy in different orchids of commercial relevance. Full article

(This article belongs to the Special Issue Plant Regeneration and Organ Formation)

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