scholarly journals Effect of Enzyme Treatments on Protoplast Isolation from Leaves of Vetiver (Vetiveria spp.)

2021 ◽  
Vol 20 (3) ◽  
Author(s):  
Sumana Leaungthitikanchana ◽  
Khachapohn Thongdonyod ◽  
Nootjaree Singphan
Keyword(s):  
Cell Wall ◽  
Protoplast Fusion ◽  
Incubation Time ◽  
Protoplast Culture ◽  
Protoplast Isolation ◽  
Fusion Technology ◽  
Vetiveria Zizanioides ◽  
Enzyme Solution ◽  
Protoplast Yield ◽  
High Viability

Protoplast isolation is a first and important step for establishing a new plant with desired traits through protoplast fusion technology. This experiments were conducted to evaluate various concentration of enzymes and incubation time on protoplast yield and viability in two vetiver ecotypes, Kamphaeng Phet 2 (Vetiveria zizanioides Nash) and Prachuap Khiri Khan (V. nemoralis A.Camus). The results revealed that protoplast yields were significantly affected by different enzyme treatments. The highest protoplast yield (6.12x105 protoplasts/ml) and high viability (98.61%) in Kamphaeng Phet 2 was obtained through the process of cell wall digestion when treated with enzyme solution containing 0.5% (w/v) cellulase onozuka R-10 and 0.5% (w/v) macerozyme R-10 in combination. While, the optimal enzyme solution for protoplast isolation from leaves of Prachuap Khiri Khan was the combination of 1.0% (w/v) cellulase onozuka R-10 and 0.4% (w/v) macerozyme R-10, resulting in the highest yield (6.80x105 protoplasts/ml) and viability (96.56%) of protoplasts. Meanwhile, incubation time of 24 h with the optimal enzyme solution resulted in the highest protoplast yields of both ecotypes. Our findings have the potential to generate an efficient protocol to isolate the protoplast from leaves of vetiver which can be used for further research studies in protoplast culture and fusion for vetiver improvement. Keywords: Cellulase onozuka R-10, Macerozyme R-10, Protoplast isolation, Vetiver

scholarly journals Teknik Isolasi dan Kultur Protoplas Tanaman Padi

2016 ◽  
Vol 3 (2) ◽  
pp. 60 ◽  
Author(s):  
Deden Sukmadjaja ◽  
Novianti Sunarlim ◽  
Endang G. Lestari ◽  
Ika Roostika ◽  
Tintin Suharlini
Keyword(s):  
Room Temperature ◽  
Protoplast Culture ◽  
Somatic Hybridization ◽  
Sucrose Solution ◽  
Protoplast Isolation ◽  
Protoplast Regeneration ◽  
Isolation And Purification ◽  
Enzyme Solution ◽  
Wild Rice Species ◽  
Rice Genotypes

<p>Protoplast<br />fusion or somatic hybridization technology is an alternative<br />technology for production hybrids of plants that are difficult<br />to be produced by conventional methods due to their sexual<br />incompatibility. An experiment was conducted to develop<br />techniques for isolation, purification, and culture of rice<br />protoplasts of cultivar IR64 and a wild rice species (Oryza<br />officinalis). Optimization of protoplast isolation and purification<br />methods from both rice genotypes were successfully<br />done. The highest protoplast density was obtained by<br />digesting embryonic callus or stems of young seedling in an<br />enzyme solution containing of 2% cellulose, 0.1% pectolyase,<br />0.5% macerozyme, 0.5% driselase, 5 mM ES, and 13% mannitol<br />in CPW solution. The protoplast digestion was done for<br />three hours by soaking in the enzyme solution followed by<br />shaking at 50 rpm under a room temperature. Purification of<br />the protoplasts were done by separating them from plant<br />debris using a 25% sucrose solution. Protoplast regeneration<br />was not successful using although different media compositions<br />and conditions. Growth process from cell division to<br />cell aggregate was only successful on IR64 protoplast culture<br />on a medium that contained AgNO3.</p>

scholarly journals CMV-RESISTANT CULTIVAR SELECTION BY ENZYME-LIKED IMMUNOSORBENT ASSAY AND PROTOPLAST ISOLATION METHOD IN CUCUMBER.

HortScience ◽  
1992 ◽  
Vol 27 (6) ◽  
pp. 672a-672
Author(s):  
Kuen-Woo Park ◽  
Min-Jea Kim
Keyword(s):  
Cell Wall ◽  
Immunosorbent Assay ◽  
Wall Thickness ◽  
Protoplast Isolation ◽  
Elisa Test ◽  
Resistant Cultivar ◽  
Isolation Method ◽  
Resistant Cultivars ◽  
Protoplast Yield ◽  
Cultivar Selection

This experiment was carried out to select resistant cultivar to CMV in cucumber using Elisa-test and protoplast isolation method. Twenty domestic cultivars or lines and 8 European cultivars were ested for resistance by Elisa test. Among the domestic cultivars, DADAKI group was found to be susceptible and CHEONGJANG group resistant. Among all the cultivars and lines tested, a European cultivar, DALIBOR and Janghyungheukjinju Korean line were found to be highly resistant. When comparing for the protoplast yield depending upon the positions of seedlings (cotyledon, young leaf and hypocotyl), the highest protoplast yield could be obtained from cotyledons in macerozyme 0.5% + cellulose 2.0%. Protoplast yields in susceptible cultivars were higher than those from resistant cultivars. Differences in cell wall thickness between susceptible and resistant cultivars were observed.

scholarly journals Isolation of Protoplasts from Nepenthes - A Plant Carnivore

2014 ◽  
Vol 24 (1) ◽  
pp. 93-100 ◽  
Author(s):  
Jaylen Sweat ◽  
Michael S. Bodri
Keyword(s):  
Cell Wall ◽  
Cell Division ◽  
Effective Treatment ◽  
Wild Species ◽  
Plant Tissue ◽  
Protoplast Isolation ◽  
Protoplast Yield ◽  
Viable Culture

Protoplasts were isolated from the lamina of greenhouse grown Nepenthes ampullaria and the hybrid N. ‘Rokko’ Exotica in order to develop a protocol for protoplast isolation suitable for wild species of Nepenthes. Various molarities utilizing mannitol or sorbitol and different enzyme mixtures and concentrations as well as incubation times were evaluated to maximize protoplast yield and viability.  The most effective treatment, a 4 hrs incubation at 40 rpm and 25°C in a solution consisting of 0.5 M sorbitol, 5% cellulase ‘Onozuka’ R-10, 0.5% macerozyme R-10, and 0.3% pectolyase Y-23, generated 4.35 × 106 protoplasts/ gfw of which 62.1% were viable.  Culture was attempted in respect of  regeneration of the cell wall, however, no cell division was observed. Plant Tissue Cult. & Biotech. 24(1): 93-100, 2014 (June) D. O. I. http://dx.doi.org/10.3329/ptcb.v24i1.19217

Influence of enzyme solution on protoplast culture and transient gene expression in maize (Zea mays L.)

1994 ◽  
Vol 39 (1) ◽  
pp. 43-48 ◽  
Author(s):  
Brigitte Krautwig ◽  
Paul A. Lazzeri ◽  
Horst L�rz
Keyword(s):  
Gene Expression ◽  
Zea Mays ◽  
Zea Mays L ◽  
Protoplast Culture ◽  
Transient Gene Expression ◽  
Enzyme Solution

scholarly journals Inhibition of phenylpropanoid biosynthesis increases cell wall digestibility, protoplast isolation, and facilitates sustained cell division in American elm (Ulmus americana)

2012 ◽  
Vol 12 (1) ◽  
pp. 75 ◽  
Author(s):  
A Maxwell P Jones ◽  
Abhishek Chattopadhyay ◽  
Mukund Shukla ◽  
Jerzy Zoń ◽  
Praveen K Saxena
Keyword(s):  
Cell Wall ◽  
Cell Division ◽  
Protoplast Isolation ◽  
Ulmus Americana ◽  
Cell Wall Digestibility ◽  
American Elm ◽  
Phenylpropanoid Biosynthesis

Protoplast Fusion Technology- Somatic Hybridization and Cybridization

2010 ◽  
pp. 175-198 ◽  
Author(s):  
Jude W. Grosser ◽  
Milica alovi ◽  
Eliezer S. Louzada
Keyword(s):  
Protoplast Fusion ◽  
Somatic Hybridization ◽  
Fusion Technology

Protoplast fusion technology

1989 ◽  
Vol 12 (4) ◽  
pp. 157-161 ◽  
Author(s):  
Stephen Gleddie ◽  
Wilfred A. Keller
Keyword(s):  
Protoplast Fusion ◽  
Fusion Technology

FUSION AND DIVISION OF NUCLEI IN MULTINUCLEATED SOYBEAN PROTOPLASTS

1971 ◽  
Vol 13 (2) ◽  
pp. 347-353 ◽  
Author(s):  
R. A. Miller ◽  
O. L. Gamborg ◽  
W. A. Keller ◽  
K. N. Kao
Keyword(s):  
Cell Culture ◽  
Cell Wall ◽  
Protoplast Fusion ◽  
Nuclear Division ◽  
Nuclear Fusion ◽  
Complete Synchronization ◽  
Cellular Division ◽  
Nuclear Behaviour ◽  
Soybean Cell

Protoplasts were produced from a soybean cell culture by enzymatic removal of the cell wall. The protoplasts were fixed after various periods of culture. There was a large amount of protoplast fusion during formation. The nuclear behaviour during division was observed. Nuclear fusion prior to nuclear division was common. Almost complete synchronization of multinucleates was found. Various abnormalities were present in nuclear and cellular division which could have led to aneuploid production.

High-affinity phosphate absorption is independent of the root cell wall in Pisum sativum

1987 ◽  
Vol 65 (7) ◽  
pp. 1504-1508 ◽  
Author(s):  
Daniel D. Lefebvre ◽  
David T. Clarkson
Keyword(s):  
Cell Wall ◽  
Phosphate Uptake ◽  
Osmotic Shock ◽  
Protoplast Isolation ◽  
Bathing Solution ◽  
Concentration Increase ◽  
Root Cells ◽  
High Affinity ◽  
Phosphate Influx ◽  
Root Cell Wall

The high affinity for phosphate influx by pea root cells was not significantly altered by osmotic shock, plasmolysis, or the preparation of free protoplasts. The Km of normal roots was 7.74 ± 2.14 μM. The Vmax for phosphate uptake was reduced to 64% by osmotic shock and to 38 and 23% by plasmolysis and protoplast isolation, respectively. Efflux analysis indicated that there was a concentration increase for phosphate in the root free space over that of the bathing solution. This apparent binding does not appear to be involved in the uptake affinity of phosphate.

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