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Artykuły w czasopismach na temat „Protoplasts”

Autor: Grafiati

Data publikacji: 4 czerwca 2021

Data aktualizacji: 25 lipca 2025

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1

Han, Jong-Eun, Han-Sol Lee, Hyoshin Lee, Hyunwoo Cho, and So-Young Park. "Embryogenic Stem Cell Identity after Protoplast Isolation from Daucus carota and Recovery of Regeneration Ability through Protoplast Culture." International Journal of Molecular Sciences 23, no. 19 (2022): 11556. http://dx.doi.org/10.3390/ijms231911556.

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Protoplasts are single cells isolated from tissues or organs and are considered a suitable system for cell studies in plants. Embryogenic cells are totipotent stem cells, but their regeneration ability decreases or becomes lost altogether with extension of the culture period. In this study, we isolated and cultured EC-derived protoplasts (EC-pts) from carrots and compared them with non-EC-derived protoplasts (NEC-pts) with respect to their totipotency. The protoplast isolation conditions were optimized, and the EC-pts and NEC-pts were characterized by their cell size and types. Both types of p

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2

Perera, Srini C., and Peggy Ozias-Akins. "Regeneration from Sweetpotato Protoplasts and Assessment of Growth Conditions for Flow-sorting of Fusion Mixtures." Journal of the American Society for Horticultural Science 116, no. 5 (1991): 917–22. http://dx.doi.org/10.21273/jashs.116.5.917.

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Petiole protoplasts of the sweetpotato [Ipomoea batatas (L.) Lam.] cultivars Red Jewel and Georgia Jet formed cell walls within 24 hours and divided in 2 to 3 days. Pretreating enzyme solutions with activated charcoal increased the viability and division frequency of protoplasts. Culture of protoplast-donor plants in a medium containing STS did not affect plant growth, protoplasm yield, or viability, but did increase the division frequency. Culture of protoplasts for 24 hours in a medium containing DB, a cell wall synthesis inhibitor, or staining of protoplasts with FDA did not significantly a

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3

Struck, C., R. Rohringer, and R. Heitefuss. "Isolation and lectin-binding properties of barley epidermal and mesophyll protoplasts." Canadian Journal of Botany 72, no. 11 (1994): 1688–91. http://dx.doi.org/10.1139/b94-207.

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Protoplasts from primary leaves of barley (Hordeum vulgare L.) were obtained by enzymatic digestion and fractionated by discontinuous density gradient centrifugation to yield highly enriched fractions of mesophyll and epidermal protoplasts. A characterization of both protoplast types resulted in a clear differentiation of the outer protoplast surfaces. The protoplasts were examined for affinity to various lectins by agglutination tests and by labeling with lectin – fluorescein isothiocyanate conjugates. Both types of protoplasts agglutinated with soybean lectin. Fluorescein isothiocyanate-labe

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4

Sinha, Anupam, Andrew C. Wetten, and P. D. S. Caligari. "Effect of biotic factors on the isolation of Lupinus albus protoplasts." Australian Journal of Botany 51, no. 1 (2003): 103. http://dx.doi.org/10.1071/bt01104.

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Several tissue types of Lupinus albus L. were investigated as sources for the isolation of protoplasts. Cotyledons from in vitro seedlings were found to yield the highest number of protoplasts compared with leaves, hypocotyls and roots. A combination of the protoplast isolation enzymes, cellulase and Pectolyase Y23, was capable of releasing the highest number of protoplasts compared with a combination of cellulase and Macerase. Protoplast yield increased with increasing cotyledon age but was accompanied by a progressive decline in protoplast viability. The optimal combination of protoplast yie

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5

Qiu, Quan-Sheng, Ze-Zhou Wang, Nang Zhang, Qi-Gui Cai, and Rong-Xi Jiang. "Aquaporins in the plasma membrane of leaf callus protoplasts of Actinidia deliciosa var. deliciosa cv. Hayward." Functional Plant Biology 27, no. 1 (2000): 71. http://dx.doi.org/10.1071/pp99033.

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The water transport activity of Actinidia deliciosa protoplasts was determined using a cell imaging system. Results showed that the protoplast volume increased swiftly when placed in a hypoton-ic medium, and also increased with an increase in medium osmotic gradients. The osmotic water permeability coefficient (Pf) values were 0.118 × 10–3, 0.121 × 10–3, and 0.133 × 10–3 cm s–1 when the osmotic gradients were 75, 100, and 125 mosmol, respectively. The water transport activity of protoplas-ts could be inhibited by HgCl2 and stimulated by amphotericin B. Moreover, ZnCl2 and ZnSO4 had a significa

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6

Ishikawa, Francine Hiromi, Quélen de Lima Barcelos, Elaine Aparecida de Souza, and Eustáquio Souza Dias. "Factors affecting the production and regeneration of protoplasts from Colletotrichum lindemuthianum." Ciência e Agrotecnologia 34, no. 1 (2010): 74–79. http://dx.doi.org/10.1590/s1413-70542010000100009.

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The present work reports factors affecting the production and regeneration of protoplasts from Colletotrichum lindemuthianum. The usefulness of protoplast isolation is relevant for many different applications and has been principally used in procedures involving genetic manipulation. Osmotic stabilizers, lytic enzymes, incubation time and mycelial age were evaluated in terms of their effects on protoplast yield. The optimal condition for protoplast production included the incubation of young mycelia (48 h) in 0.6 mol l-1 NaCl as the osmotic stabilizer, with 30 mg ml-1 Lysing Enzymes from Trich

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7

Leinhos, Volker, and Rodney Arthur Savidge. "Isolation of protoplasts from developing xylem of Pinusbanksiana and Pinusstrobus." Canadian Journal of Forest Research 23, no. 3 (1993): 343–48. http://dx.doi.org/10.1139/x93-050.

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Protoplasts were isolated from developing xylem of Pinusbanksiana Lamb, and Pinusstrobus L. by incubating freshly harvested tissue in a cellulose–pectinase mixture having mannitol as osmoticum. Protoplasts were then purified using a discontinuous sucrose–mannitol gradient. More than 70% of the isolated protoplasts were of small diameter (12–27 μm) and had dense cytoplasm and many small vacuoles, suggesting that they originated from ray cells. Larger protoplasts constituted about 25% of the protoplast population; these contained single large vacuoles and only parietal cytoplasm, suggesting that

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8

Tusa, N., J. W. Grosser, and F. G. Gmitter. "Plant Regeneration of `Valencia' Sweet Orange, `Femminello' Lemon, and the Interspecific Somatic Hybrid following Protoplasm Fusion." Journal of the American Society for Horticultural Science 115, no. 6 (1990): 1043–46. http://dx.doi.org/10.21273/jashs.115.6.1043.

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Protoplasm culture following the chemical fusion of `Valencia' sweet orange [Citrus sinensis (L.) Osb.] protoplasts, isolated from an embryogenic suspension culture, with `Femminello' lemon [Citrus limon (L.) Burro. f.] leaf protoplasts resulted in the regeneration of an interspecific allotetraploid somatic hybrid plant, two autotetraploid lemon plants, and diploid plants from both parents. The regeneration of plants from lemon leaf protoplasts is an example of protoplast-to-plant regeneration from non-nucellus-derived tissue for Citrus. Regenerated plants were classified according to leaf mor

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9

Saxena, Praveen K., and John King. "Optimal conditions for the isolation and culture of protoplasts from a cell suspension culture of an isoleucine–valine-requiring auxotroph of Datura innoxia." Canadian Journal of Botany 65, no. 8 (1987): 1736–40. http://dx.doi.org/10.1139/b87-237.

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Conditions have been standardized for obtaining high yields of viable protoplasts from cell suspension cultures of an isoleucine–valine-requiring auxotroph (IV-1) of Datura innoxia P. Mill. Isolation of protoplasts critically required the use of a salt solution, containing sodium chloride and potassium chloride (0.125 M each), as osmotic stabilizers. The yield, viability, and divisional activity of the protoplasts isolated with mannitol or sucrose were poor. Protoplast-releasing enzymes used to isolate IV-1 protoplasts could be used twice without any loss in the yield or viability of the proto

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10

Liu, Donglong, and Nancy A. Reichert. "PROTOPLAST ISOLATION AND CULTURE OF KENAF (HIBISCUS CANNABINUS L.)." HortScience 29, no. 7 (1994): 729e—729. http://dx.doi.org/10.21273/hortsci.29.7.729e.

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Protoplast isolation and culture protocols were developed for leaf tissue from 6 kenaf cultivars [Everglades 41 (E41), E71, Guatemala 4 (G4), G45, G51, and Tainung 1]. For protoplast isolation, the best combination of hydrolytic enzymes was cellulysin (1% w/v; Calbiochem) plus macerase (0.5% w/v; Calbiochem), with a 24 hour digestion at 30°C in the dark. Yields reached 7.2 (10)6 protoplasts/g leaf tissue. Protoplast viabilities ranged from 65% to 96%. Minor cultivar differences were observed related to protoplast yield, but all viability estimates were in an acceptable range. Greatest cell div

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11

Riyadi, Imron. "Isolasi Protoplas Tanaman Kacang Panjang secara Enzimatis." Buletin Plasma Nutfah 12, no. 2 (2016): 62. http://dx.doi.org/10.21082/blpn.v12n2.2006.p62-68.

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<p>The technique, kind and concentration of enzyme that were appro-priate and optimum affected the isolation process and rendement result of plant protoplasts. A research was conducted to enhance the protoplast rendements of long bean (Vigna sinensis, L.) that was isolated by enzyme Cellulase RS and Macerozyme R-10 as single and combination in a solution. Concentrations of enzyme were used as much as 2.0-3.0% w/v for Cellulase RS and 0.4-0.6% w/v for Macerozyme R-10. Those solutions contain mannitol 25 mM as osmotycum. Isolation process was done on shaker with 50 rpm (rotation per minute

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12

Attree, S. M., D. I. Dunstan, and L. C. Fowke. "Initiation of embryogenic callus and suspension cultures, and improved embryo regeneration from protoplasts, of white spruce (Picea glauca)." Canadian Journal of Botany 67, no. 6 (1989): 1790–95. http://dx.doi.org/10.1139/b89-227.

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Rapid and high frequency somatic embryo regeneration from protoplasts isolated from 10 embryogenic cell lines of white spruce (Picea glauca) is reported. Embryogenic callus was initiated from immature zygotic embryos as source material for protoplast isolation. Individual cell lines exhibited different capabilities for sustained growth. Protoplast plating efficiencies depended on the concentrations of macroelements included in the medium. Using a medium with reduced salts, individual protoplasts developed directly into embryos with no disorganized growth period. Protoplasts from newly establis

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13

Moon, Ki-Beom, Ji-Sun Park, Su-Jin Park, et al. "A More Accessible, Time-Saving, and Efficient Method for In Vitro Plant Regeneration from Potato Protoplasts." Plants 10, no. 4 (2021): 781. http://dx.doi.org/10.3390/plants10040781.

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Both obtaining high-yielding, viable protoplasts and following reliable regeneration protocols are prerequisites for the continuous expansion and development of newly emerging systems involving protoplast utilization. This study determines an efficient process from protoplast isolation to shoot regeneration in vitro. The maximum yield of protoplast extraction, which was 6.36 ± 0.51 × 106 protoplasts/g fresh weight (FW), was approximately 3.7 times higher than that previously reported for potato protoplasts. To obtain data, wounded leaves were used by partially cutting both sides of the midrib,

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14

Stajič, Ester. "Improvements in Protoplast Isolation Protocol and Regeneration of Different Cabbage (Brassica oleracea var. capitata L.) Cultivars." Plants 12, no. 17 (2023): 3074. http://dx.doi.org/10.3390/plants12173074.

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Protoplasts are a versatile tool in plant biotechnology since they can be used for basic biological studies as well as for breeding strategies based on genome editing. An efficient protoplast isolation protocol is essential for conducting protoplast-based studies. To optimize the protoplast isolation protocol in cabbage (Brassica oleracea var. capitata L.), different enzyme solutions were tested for the isolation of leaf mesophyll protoplasts. In our experiments, the combination of 0.5% Cellulase Onozuka RS and 0.1% Macerozyme R-10 showed the best result. The optimized protocol proved suitable

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15

Tran, Huong Thanh, and Cuong Quoc Vo. "Protoplast isolation and culture from different explants of Musa spp. Cau man." Science and Technology Development Journal - Natural Sciences 1, no. 6 (2018): 106–16. http://dx.doi.org/10.32508/stdjns.v1i6.621.

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In this paper, the roles of type and concentration of enzymes on protoplast isolation from in vitro leaves, multi-scalps (highly proliferating meristem culture), and young male flower of banana cv. cau man were studied. Respiration rate and content of plant hormones of these materials were analysed. Different techniques were used to culture these protoplasts. The development of protoplasts was observed under fluorescence microscope. The highest yield of protoplast (69.5 x 106 protoplasts / g fresh weight) was obtained from young male flowers after 16 hours treatment with 1.5 % cellulase, 0.25

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16

Li, Cai Hong, De Feng Xu, Ri Ying Ye, Ya Ling Wang, and Li Jun Sun. "An Efficient Method for Monitoring and Improving the Regeneration Efficiency of Protoplasts from Aspergillus Oryzae HN3042." Advanced Materials Research 781-784 (September 2013): 808–11. http://dx.doi.org/10.4028/www.scientific.net/amr.781-784.808.

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Protoplast technique is a powerful tool for strain improvement and the preparation of protoplasts with high regeneration efficiency is the basis of subsequent manipulations. In this study, the effects of enzymatic hydrolysis time on release and regeneration of protoplasts from Aspergillus oryzae HN3042 were investigated by analyzing the time-series image. The results showed that there was a significant correlationship between regeneration efficiency and morphological size of protoplasts (P < 0.01). An efficient method for monitoring and improving the regeneration efficiency of protoplast wa

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17

Pauls, K. P., and P. V. Chuong. "Flow cytometric identification of Brassica napus protoplast fusion products." Canadian Journal of Botany 65, no. 5 (1987): 834–38. http://dx.doi.org/10.1139/b87-113.

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A flow cytometric method to identify Brassica somatic hybrids has been developed. The procedure is based on the use of fluorescein isothiocyanate stained hypocotyl protoplasts and unstained mesophyll protoplasts as partners for polyethylene glycol induced protoplast fusion. The fluorescein isothiocyanate stained hypocotyl protoplasts and unstained mesophyll protoplasts were passed through the flow cytometer – cell sorter separately to maximize the sensitivity of the red and green detectors to chlorophyll and fluorescein isothiocyanate fluorescence, respectively, and to ensure that there was no

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18

Kurita-Tashiro, Asami, Noriko Hayashi, Tomoya Oyanagi, and Hamako Sasamoto. "New Factors for Protoplast-Callose-Fiber Formation in Salt-Tolerant Mangrove Plants, Avicennia alba and Bruguiera sexangula and Analysis of Fiber Substructures." Journal of Plant Studies 9, no. 2 (2020): 1. http://dx.doi.org/10.5539/jps.v9n2p1.

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Elongated and spiral &beta;-1,3-glucan (callose) fibers were obtained by new factors from protoplasts cultured in liquid medium from suspension cultured cells of two salt-tolerant mangrove species; Avicennia alba and Bruguiera sexangula. Differences in salt factor for protoplast-fiber formation were compared with those of the callose fibers developed from protoplasts of non-mangrove tree plants, Larix leptolepis and Betula platyphylla, which high concentrations of divalent cations, Mg2+ (50 mM) or Ca2+ (100 mM), were stimulatory. In the halophilic A. alba protoplasts, whose cell division w

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Kantharajah, AS, and WA Dodd. "Factors That Influence the Yield and Viability of Cucumber (Cucumis sativus L) Cotyledon Protoplasts." Australian Journal of Botany 38, no. 2 (1990): 169. http://dx.doi.org/10.1071/bt9900169.

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Protoplasts isolated from cotyledons of aseptically germinated cucumber seedlings were divided into three size classes. The relationships between tissue age, isolation procedure, yield and protoplast size were investigated. During germination and up to an age of 13 days, the percentage of protoplasts in each size class underwent considerable change with a big reduction in percentage of the largest protoplasts in older cotyledons. Protoplast size and yield could also be manipulated by varying the isolation technique. In this context, temperature, incubation time and shaker speed were significan

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Sasamoto, Hamako, Tsukasa Iwashina, Sakae Suzuki, Yoshitaka Azumi, and Yoshiharu Fujii. "Evaluation of an Anthocyanin, Cyanidin 3,5-di-O-glucoside, as an Allelochemical in Red Callus of a Mangrove Sonneratia ovata, Using Protoplast Co-Culture Bioassay Method with Digital Image Analysis." Journal of Plant Studies 7, no. 2 (2018): 1. http://dx.doi.org/10.5539/jps.v7n2p1.

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protoplasts to examine the allelopathic activities. Protoplasts were isolated with Cellulase R10 and Driselase 20 in 0.6 M mannitol solution and purified by density gradient centrifugation on 0.6 M sucrose. Protoplasts were co-cultured in 50 μL of liquid Murashige and Skoog’s (MS) basal medium containing 1 μM 2,4-dichlorophenoxyacetic acid and 0.1 μM benzyladenine and 0.6 M mannitol solution in a 96-well culture plate. Protoplast density ranged from 5 × 103/mL to 105/mL. Cell division of lettuce protoplasts was strongly inhibited by addition of S. ovata protoplasts, and non-spherical cell enla

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Al-Nema, Qutaiba, and Mozahim AL-Mallah. "Electrofusion of mesophyll protoplasts from two varieties of sugar beet, (Beta vulgaris L.)." Journal of Life and Bio Sciences Research 1, no. 1 (2020): 22–25. http://dx.doi.org/10.38094/jlbsr117.

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Somatic hybridization between different plants through protoplast fusion represent an efficient experimental approach to produce genetically transformed plant species. Electrofution of mesophyll protoplasts in sugar beet was occurred to overcome the barriers faced breeding program of this economically industrial crop Protoplasts were successfully isolated from leave's mesophyll of two varieties of sugar beet (Beta vulgaris L.). Various enzyme solutions were assessed for the cell wall degrading ability. They express different efficiency in isolation of mesophyll protoplasts of var. Baraka. The

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Meng, Ruirui, Chenchen Wang, Lihua Wang, et al. "An efficient sorghum protoplast assay for transient gene expression and gene editing by CRISPR/Cas9." PeerJ 8 (October 13, 2020): e10077. http://dx.doi.org/10.7717/peerj.10077.

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Protoplasts are commonly used in genetic and breeding research. In this study, the isolation of sorghum protoplasts was optimized and applied to transient gene expression and editing by CRISPR/Cas9. The protoplast was most viable in 0.5 M mannitol, which was the highest of three concentrations after 48- and 72-hours treatments. Using this method we can derive an average of 1.6×106 cells which vary from 5 to 22 nm in size. The average transfection of the protoplasts was 68.5% using the PEG-mediated method. The subcellular assays located Sobic.002G279100-GFP and GFP proteins in the cell compartm

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Inoue, Aya, Daisuke Mori, Reiko Minagawa, Yoshiharu Fujii, and Hamako Sasamoto. "Allelopathy in a Leguminous Mangrove Plant, Derris indica: Protoplast Co-culture Bioassay and Rotenone Effect." Natural Product Communications 10, no. 5 (2015): 1934578X1501000. http://dx.doi.org/10.1177/1934578x1501000512.

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To investigate allelopathic activity of a leguminous mangrove plant, Derris indica, the ‘Protoplasts Co-culture Method’ for bioassay of allelopathy was developed using suspension culture. A suspension culture was induced from immature seed and sub-cultured in Murashige and Skoog's (MS) basal medium containing 10 μM each of 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzyladenine (BA). The protoplasts were isolated using the separate wells method with 2% each of Cellulase RS, Driselase 20 and Macerozyme R10 in 0.4 M mannitol solution. Protoplast cultures of D. indica revealed that high concen

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Wang, Lun, Jiaojiao Zhang, and Xiaoyong Xu. "A Comparison of DNA-Methylation during Protoplast Culture of Ponkan Mandarin (Citrus reticulata Blanco) and Tobacco (Nicotiana tabacum L.)." Plants 13, no. 20 (2024): 2878. http://dx.doi.org/10.3390/plants13202878.

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The epigenetic variation in protoplast regeneration is a topic that has attracted interest recently. To elucidate the role of DNA methylation in the regeneration of protoplasts from the ponkan (Citrus reticulata), this study employs the methylation-sensitive amplification polymorphism (MSAP) molecular marker technique to analyze changes in DNA methylation levels and patterns during the isolation and culture of protoplasts from ponkan and tobacco. Additionally, differential DNA methylation fragments are cloned, sequenced, and subjected to bioinformatics analysis. The results reveal that, for no

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Jenes, Barnabas, Matti Puolimatka, Pedro Bittencourt, and Seppo Pulli. "Time saving method for protoplast isolation, transformation and transient gene expression assay in barley." Agricultural and Food Science 3, no. 2 (1994): 199–205. http://dx.doi.org/10.23986/afsci.72695.

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This study was conducted to establish a rapid method for barley (Hordeum vulgare L.) protoplast isolation to provide an easy-to-use procedure for the transformation and primary investigation of new gene constructs by transient gene expression assays. Protoplasts were successfully isolated from the chopped embryo and scutellum parts of mature barley seeds by digesting three hours with an enzyme mixture. Isolated protoplasts were washed in W5 washing solution, sieved through plastic meshes and then cleaned on sucrose gradient. The suitability of these directly from embryo-scutellum complexes der

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Wang, Peilin, Yuanchun Pu, Muhammad Ali Abid, et al. "A Rapid and Efficient Method for Isolation and Transformation of Cotton Callus Protoplast." International Journal of Molecular Sciences 23, no. 15 (2022): 8368. http://dx.doi.org/10.3390/ijms23158368.

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Protoplasts, which lack cell walls, are ideal research materials for genetic engineering. They are commonly employed in fusion (they can be used for more distant somatic cell fusion to obtain somatic hybrids), genetic transformation, plant regeneration, and other applications. Cotton is grown throughout the world and is the most economically important crop globally. It is therefore critical to study successful extraction and transformation efficiency of cotton protoplasts. In the present study, a cotton callus protoplast extraction method was tested to optimize the ratio of enzymes (cellulase,

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Sun, M., H. Kieft, and AAM van Lammeren. "Cotyledon-derived diploid and haploid protoplast culture and diploid plant regeneration in Brassica napus cv. ' Topas '." Canadian Journal of Botany 76, no. 3 (1998): 530–41. http://dx.doi.org/10.1139/b98-022.

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The present paper describes a simple and reliable protocol for the successful isolation, purification, culture, and regeneration of diploid cotyledon-derived protoplasts of Brassica napus L. cv. 'Topas'. Various protoplast isolation media, nutrient media, subculture procedures, and protoplast sources were tested under two culture temperatures. Protoplast viability, cell wall regeneration, and cell division were monitored. Single cotyledon-derived protoplasts formed calli in liquid protoplast medium, and when these were subcultured on solid proliferation medium and solid regeneration medium of

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Schulz, Margot, and Gottfried Weissenböck. "Isolation and Separation of Epidermal and Mesophyll Protoplasts from Rye Primary Leaves — Tissue-Specific Characteristics of Secondary Phenolic Product Accumulation." Zeitschrift für Naturforschung C 41, no. 1-2 (1986): 22–27. http://dx.doi.org/10.1515/znc-1986-1-205.

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Abstract We have develop ed a technique for the large-scale isolation of epidermal and mesophyll protoplasts, as well as the vascular strands, of rye primary leaf blades. Separation of the two types of protoplasts has been successful only from leaves harvested at the end of a 13-h light period, when chloroplasts were enriched in starch. The occurrence of different flavonoid compounds, and amounts, in epidermal and mesophyll protoplasts can be used as criteria for protoplast purity and viability since C-glucosylflavone O-glycosides are characteristic of epidermal protoplasts whereas flavone O-

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Mieth, Hannelore, Volker Speth, and Jürgen Ebel. "Phytoalexin Production by Isolated Soybean Protoplasts." Zeitschrift für Naturforschung C 41, no. 1-2 (1986): 193–201. http://dx.doi.org/10.1515/znc-1986-1-229.

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Abstract Protoplasts isolated enzymatically from suspension-cultured cells of soybean (Glycine max L. Merr., cv. Harosoy 63) were used to study the production of the isoflavonoid-derived phytoalexin, glyceollin. A large enhancement in the in vivo rates of synthesis and catalytic activities of two of the enzymes associated with glyceollin biosynthesis, phenylalanine ammonia-lyase and chalcone synthase, preceded phytoalexin accumulation during early stages of culture of isolated protoplasts while cell wall regeneration occurred. A glucan elicitor from cell walls of the fungus Phytophthora megas

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Sasamoto, Hamako, and Shinso Yokota. "In vitro Bioassay of Allelopathic Activities of a Mangrove Tree, Kandelia obovata, and Fast-growing Trees, Betula platyphylla and Populus alba, Using Protoplast Co-culture Method." Journal of Plant Studies 10, no. 2 (2021): 8. http://dx.doi.org/10.5539/jps.v10n2p8.

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Allelopathic activities of a salt-tolerant and low-temperature tolerant mangrove tree, Kandelia obovata, which grows in brackish water regions of sub-tropical areas, and two fast-growing trees, Betula platyphylla and Populus alba, which grow in the temperate area, were examined by two in vitro bioassay methods, the sandwich method using dried leaves and the protoplast co-culture method using leaf protoplasts. Lettuce root growth examined by the sandwich method, was inhibited 50% by 50 mg dried mature leaves of K. obovata. In the protoplast co-culture method, inhibition rates of cell division o

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Echeverria, Ed. "Preparation and Characterization of Protoplasts from Citris Juice Vesicles." Journal of the American Society for Horticultural Science 112, no. 2 (1987): 393–96. http://dx.doi.org/10.21273/jashs.112.2.393.

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Abstract A method for the preparation of protoplasts from juice vesicles of citrus fruits is described. Protoplasts were obtained after incubating juice vesicles for 16 hr at room temperature in a medium containing cellulase and pectinase. The solution containing the protoplast was filtered through a 200-μm nylon mesh and the filtrate layered on a Percoll density gradient of 4%, 3.5%, and 3%. After 1 to 2 hr at 4°C, protoplasts were collected from the 4% Percoll layer. Protoplast viability was assessed by different methods and estimated to be ≈ 85%. The use of several stains and observations w

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32

Bekkaoui, Faouzi, and David I. Dunstan. "Permeabilization of Piceaglauca protoplasts to macromolecules." Canadian Journal of Forest Research 19, no. 10 (1989): 1316–21. http://dx.doi.org/10.1139/x89-202.

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Chemical permeabilization (polyethylene glycol, molecular weight 3350) and electropermeabilization (electroporation) treatments were applied to white spruce protoplasts to determine their effectiveness for uptake of membrane impermeable macromolecules. The two techniques have been compared using the membrane impermeable fluorescent dye calcein (molecular weight 622). The effects of varying the polyethylene glycol concentration, or the capacitance and voltage, were tested. In both techniques, the viability of protoplasts decreased after treatment compared with the controls. However, electropora

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33

Shao, Yingying, Detian Mu, Limei Pan, et al. "Optimization of Isolation and Transformation of Protoplasts from Uncaria rhynchophylla and Its Application to Transient Gene Expression Analysis." International Journal of Molecular Sciences 24, no. 4 (2023): 3633. http://dx.doi.org/10.3390/ijms24043633.

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Protoplast-based engineering has become an important tool for basic plant molecular biology research and developing genome-edited crops. Uncaria rhynchophylla is a traditional Chinese medicinal plant with a variety of pharmaceutically important indole alkaloids. In this study, an optimized protocol for U. rhynchophylla protoplast isolation, purification, and transient gene expression was developed. The best protoplast separation protocol was found to be 0.8 M D-mannitol, 1.25% Cellulase R-10, and 0.6% Macerozyme R-10 enzymolysis for 5 h at 26 °C in the dark with constant oscillation at 40 rpm/

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34

Krasova, Yuliya V., Vladimir V. Fadeev, Yelizaveta М. Moiseeva, Yury S. Gusev, and Mikhail I. Chumakov. "Optimization of the technique for maize protoplast isolation and their nativity after electroporation." Izvestiya of Saratov University. Chemistry. Biology. Ecology 22, no. 4 (2022): 445–54. http://dx.doi.org/10.18500/1816-9775-2022-22-4-445-454.

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We optimizes the composition and concentration of the enzymes, the time for enzymatic treatment and the volume of the enzyme mixture. We also optimized the concentration osmotic of agent, the centrifugation mode, and filter pore size for protoplasts isolating from epidermal cells of maize roots (Zea mays L.) of the Brown Marker (BM) line. It was found that 150 minutes is the optimal time for 150 mg root tissue maceration. The yield of intact protoplasts was ~ 4.4 ± 0.2 × 105 cells/mL at the following concentrations of enzymes and osmotic stabilizer: cellulase – 17,4, pectolase – 1.2, hemicellu

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35

Balasubramanian, N., G. Annie Juliet, P. Srikalaivani, and D. Lalithakumari. "Release and regeneration of protoplasts from the fungus Trichothecium roseum." Canadian Journal of Microbiology 49, no. 4 (2003): 263–68. http://dx.doi.org/10.1139/w03-034.

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A protocol for isolating and regenerating protoplasts from Trichothecium roseum has been described. Protoplasts from T. roseum were isolated using (i) a lytic enzyme combination composed of Novozym 234, chitinase, cellulase, and pectinase at a 5-mg/mL concentration and (ii) 0.6 M KCl as an osmotic stabilizer. A maximum number of 28 × 104 protoplasts/mL were obtained at pH 5.5. Experiments on the regeneration and reversion of protoplasts revealed a maximum regeneration (60.8%) in complete medium (potato dextrose yeast extract agar) amended with 0.6 M KCl. The regenerated protoplasts were simi

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36

Li, Xiaobin, Ying Qin, Yufei Kong, Samantha Chandranath Karunarathna, Yunjiang Liang, and Jize Xu. "Optimization of Protoplast Preparation Conditions in Lyophyllum decastes and Transcriptomic Analysis Throughout the Process." Journal of Fungi 10, no. 12 (2024): 886. https://doi.org/10.3390/jof10120886.

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Protoplasts are essential tools for genetic manipulation and functional genomics research in fungi. This study systematically optimized protoplast preparation conditions and examined transcriptional changes throughout the preparation and regeneration processes to elucidate the molecular mechanisms underlying the formation and regeneration of protoplasts in Lyophyllum decastes. The results indicated an optimal protoplast yield of 5.475 × 106 cells/mL under conditions of fungal age at 10 days, digestion time of 2.25 h, enzyme concentration of 2%, and digestion temperature of 28 °C. The Z5 medium

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37

Sun, Yuan, Zheng, et al. "An Efficient and Economical Protocol for Isolating, Purifying and PEG-Mediated Transient Gene Expression of Chinese Kale Hypocotyl Protoplasts." Plants 8, no. 10 (2019): 385. http://dx.doi.org/10.3390/plants8100385.

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In this study, we report the isolation and purification of protoplasts from Chinese kale (Brassica oleracea var. alboglabra) hypocotyls, and their transient gene expression transformation and subcellular localization of BaMYB75 (Bol042409). The upshot is that the vintage protocol included 5-d hypocotyls that were enzymatically hydrolyzed for 8 h in enzyme solution (3.0% cellulase, 0.5% pectolase, and 0.5 M mannitol), and the protoplasts were purified by precipitation. The total yield of protoplasts was 8 × 105 protoplast g−1 fresh weight, and the protoplasts’ viability was 90%. The maximum tra

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38

Al-Ne'ma, Q. Sh, and M. K. Al-Mallah. "Protoplast isolation from leaf mesophyll of sugarbeet Beta vulgaris L. axenic seedlings." Journal of Biotechnology Research Center 7, no. 3 (2013): 36–42. http://dx.doi.org/10.24126/jobrc.2013.7.3.280.

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Protoplasts were isolated from leaf mesophyll of sugarbeet (Beta vulgaris L.) axenic seedlings. Eight enzyme mixtures were tested for cell wall degrading ability. The efficient enzyme solution was mixture "II" that consist of 1.5% Cellulase RS, 2% Cellulase R10, 1% Macerozym R10 and 0.1% Pectolyase Y23. This mixture was efficient in releasing protoplasts and gave high yield of a density 7.3 × 104 protoplast / ml. These isolated protoplasts were viable 93%, their sizes ranged from 13 up to 52 µm, vacuolated and unvacuolated. This finding enable workers to focus on somatic hybridization through

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39

Martin, F. R., and R. A. Nolan. "American cockroach (Periplaneta americana) hemocyte response to Entomophaga aulicae protoplasts." Canadian Journal of Zoology 64, no. 6 (1986): 1369–72. http://dx.doi.org/10.1139/z86-204.

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Contact through protoplasmic extensions from the protoplasts was infrequently observed and cell–cell contact involving large surface areas was not observed when adult Periplaneta americana hemocytes were incubated with Entomophaga aulicae protoplasts in in vitro culture for up to 30 min. Such contact did not appear to alter the subsequent morphology of either the protoplast or the hemocyte. Coagulocyte, plasmatocyte, and granulocyte aggregates along with individual coagulocytes and granulocytes produced a melaninlike material upon incubation with protoplasts. Incubation of the hemocytes in the

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40

Du, Jiageng, Huitao Zhang, Weilong Li, et al. "Optimization of Protoplast Preparation System from Leaves and Establishment of a Transient Transformation System in Apium graveolens." Agronomy 13, no. 8 (2023): 2154. http://dx.doi.org/10.3390/agronomy13082154.

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Protoplast culture and transformation technology offer a novel method for developing new plant varieties. Nonetheless, the effective preparation of protoplasts and transformation technology specific to celery has yet to be achieved. This study utilized celery seedling leaves as the primary materials to examine the key factors influencing protoplast isolation. The aim was to prepare leaf protoplasts with a high yield and of high quality and subsequently conduct transient gene transformation and expression. The findings indicated that the most effective procedure for isolating and purifying prot

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41

KESKITALO, M. "Can protoplast production from in vitro cultured shoots of Tanacetum vary during the season?" Agricultural and Food Science 10, no. 3 (2001): 145–51. http://dx.doi.org/10.23986/afsci.5688.

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Two different experiments were carried out to study the production of protoplasts and the variation of protoplast yield from in vitro cultured shoot tips of tansy (Tanacetum vulgare L.) and pyrethrum (Tanacetum cinerariifolium (Trevir.) Schiltz-Bip). In the first experiment, light had more pronouced effect for tansy than for pyrethrum. When the donor tissues of tansy were cultured under high light intensity the leaves contained anthocyanin and became brown during enzyme maceration. In contrast, donor tissues cultured under low light intensity produced leaves without anthocyanin. Depending on t

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42

Wei, Xiuyan, Xinyue Song, Dong Dong, et al. "Efficient production of Aschersonia placenta protoplasts for transformation using optimization algorithms." Canadian Journal of Microbiology 62, no. 7 (2016): 579–87. http://dx.doi.org/10.1139/cjm-2015-0770.

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The insect pathogenic fungus Aschersonia placenta is a highly effective pathogen of whiteflies and scale insects. However, few genetic tools are currently available for studying this organism. Here we report on the conditions for the production of transformable A. placenta protoplasts using an optimized protocol based on the response surface method (RSM). Critical parameters for protoplast production were modelled by using a Box–Behnken design (BBD) involving 3 levels of 3 variables that was subsequently tested to verify its ability to predict protoplast production (R2 = 0.9465). The optimized

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43

Kropp, Bradley R., and J. A. Fortin. "Formation and regeneration of protoplasts from the ectomycorrhizal basidiomycete Laccaria bicolor." Canadian Journal of Botany 64, no. 6 (1986): 1224–26. http://dx.doi.org/10.1139/b86-167.

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Protoplasts were released from dikaryotic mycelium of the ectomycorrhizal basidiomycete Laccaria bicolor using the lytic enzyme preparation NovoZyme 234. Protoplast release depended strongly on mycelium age, osmotic stabilizer, and temperature. The protoplasts could regenerate to form both monokaryotic and dikaryotic cultures capable of forming normal ectomycorrhizae with Pinus banksiana.

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44

Kiełkowska, Agnieszka, and Adela Adamus. "Embedding in Filter-Sterilized Alginate Enhances Brassica Oleracea L. Protoplast Culture." Acta Biologica Cracoviensia s. Botanica 56, no. 2 (2015): 20–26. http://dx.doi.org/10.2478/abcsb-2014-0018.

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Abstract The influence of sodium alginate sterilization on the viability and mitotic activity of embedded protoplasts was studied in protoplasts of Brassica oleracea subsp. alba and rubra isolated from hypocotyl tissue and leaves of seedlings or plants grown in vitro. Both leaf and hypocotyl-derived protoplasts were more viable and divided more frequently when embedded in filtrated alginate. Division frequency was highest in cv. Reball F1 and the mitotic activity of its protoplasts was three times higher when embedded in filtrated alginate (36.1 ± 6.8%) than when cultured in autoclaved alginat

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45

Jażdżewska, Elżbieta, Aleksandra Niklas, and Anna Majewska-Sawka. "Progress towards sugar beet improvement through somatic hybridization I. Inactivation of nuclei and cytoplasm in donor and recipient protoplasts." Acta Societatis Botanicorum Poloniae 64, no. 4 (2014): 341–47. http://dx.doi.org/10.5586/asbp.1995.044.

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The isolation and culture of suspension-derived protoplasts from two sugar beet (<em>Beta vulgaris</em> L.) genotypes are described. Immobilization of protoplasts in agarose resulted in high frequency divisions and microcallus regeneration, with plating efficiency (PE) being clearly genotype-dependent. In further studies towards asymmetric fusion experiments, the effect of different doses of ultraviolet radiation (UV) and iodoacetic acid (IA) on protoplast physiology was assessed. Viability of both treated (UV, IA) and untreated protoplasts (control) was determined by FDA staining,

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46

Nolan, Richard A. "Stage-specific changes in cytoplasmic protein synthesis in Entomophaga aulicae protoplasts." Canadian Journal of Microbiology 35, no. 3 (1989): 373–78. http://dx.doi.org/10.1139/m89-057.

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The patterns of protein synthesis associated with three sequential stages in protoplast morphogenesis (spindle-shaped, early fusion sphere, and late fusion sphere protoplasts) of the fungus Entomophaga aulicae were studied using both one-dimensional gels with general protein staining and two-dimensional gels with [35S]methionine protein labelling and fluorography. A total of 332 proteins were observed with 63.5% (211) common to all three developmental stages. Of the individual totals, 3.3% (8 out of 245), 7.3% (22 out of 301), and 4.5% (13 out of 286) of the proteins were unique to the spindle

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47

Sihachakr, Darasinh, and Georges Ducreux. "Isolement et culture de protoplastes à partir de petioles et de tiges de deux variétés de patate douce (Ipomoea batatas)." Canadian Journal of Botany 65, no. 1 (1987): 192–97. http://dx.doi.org/10.1139/b87-025.

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Protoplasts isolated from petioles and stems of two varieties of sweet potato showed high plating efficiency, and rhizogenic calli were obtained. The yield of protoplasts and the rate of survival of cell division were statistically analysed. Results are discussed in relation to the identification of different protoplast subpopulations and their histological origin.

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48

Stevenson, I. "Protoplast Formation from Submerged Mycelium and from Spore Germinants of Streptomyces coelicolor." Australian Journal of Biological Sciences 38, no. 1 (1985): 175. http://dx.doi.org/10.1071/bi9850175.

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Stages in the formation of protoplasts from S. coelicolor strain A3(2) have been studied by transmission electron microscopy. Protoplasts liberated from submerged mycelial growth were variable in size and were released when digestion of the cell wall by lysozyme had completely or almost completely taken place. Protoplasts did not fully adopt the typical rounded shape until after release. A single region of cytoplasm gave rise to more than one protoplast unit. Protoplasts released from spore germinants escaped from the tip of the germ tube, which was the region of the cell wall most susceptible

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49

Taylor, PWJ, HL Ko, and SW Adkins. "Factors Affecting Protoplast Isolation and the Regeneration of Shoot-Like Structures From Protoplast-Derived Callus of Sugarcane (Saccharum spp Hybrids)." Australian Journal of Botany 40, no. 6 (1992): 863. http://dx.doi.org/10.1071/bt9920863.

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High yields of protoplasts were isolated from non-regenerable, homogeneous cell suspension cultures of sugarcane, compared with regenerable, heterogeneous cell suspension cultures after incubation in an enzyme composition containing Cellulase RS, Pectinase, Macerozyme and Driselase. Higher yields of protoplasts were released from heterogeneous cell suspension cultures after the addition of 10 mg L-1 silver nitrate to the culture medium; however, ethylene production was not involved in protoplast isolation. Use of 0-05-2% Pectolyase Y23 pectinase rather than other pectinases resulted in higher

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50

Chen, Chi-Chang, Feng-Ming Lee, and Yen-Yu Kao. "Endoreduplication in leaf protoplasts of haploid Nicotiana plumbaginifolia cultured in vitro." Genome 30, no. 5 (1988): 615–20. http://dx.doi.org/10.1139/g88-104.

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When protoplasts isolated from the leaves of haploid plants of Nicotiana plumbaginifolia Viviani were examined cytologically during in vitro culture, a small fraction was found to contain diplochromosomes at the first mitosis. DNA measurements of freshly isolated protoplasts revealed that 92% contained G1 and 8% contained G2 haploid nuclei. The origin of the diplochromosomes was investigated by a simplified sister chromatid differential staining technique and autoradiography. Our results showed that nuclear division in the G1 protoplasts was normal, with each round of DNA replication being fol

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