Topic
Protoplast
About: Protoplast is a research topic. Over the lifetime, 5474 publications have been published within this topic receiving 122468 citations.
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01 Jan 1987
TL;DR: A method was developed for culturing protoplasts freshly isolated from developing soybean (Glycine max L.) cotyledons using a polyethylene glycol-electroporation technique, and CAT activity was detected in protoplast cultures 6 hours after introduction of a 35S-CAT-nopaline synthase 3' chimeric gene.
Abstract: Amethod wasdeveloped forculturing protoplasts freshly isolated from developing soybean (Glycine maxL.)cotyledons. First cell divisions were observed within 5days after protoplast isolation andmicrocalli, consisting ofabout 20cells, wereformed within 10days. Thirty days after protoplast isolation, callus tissues wereobserved without theaidofamicroscope. A 30to50%plating efficiency wasconsistently obtained. Using apolyethylene glycol-electroporation technique, DNA wasintroduced into these protoplasts. Theprotoplasts werethencultured toformcallus. Chloramphenicol acetyltransferase (CAT)activity wasdetected inprotoplast cultures 6hours after introduction ofa35S-CAT-nopaline synthase 3' chimeric gene. Thehighest CATactivity wasdetected in3-day-old electroporated protoplast cultures, indicating transient expression ofthe introduced gene. SomeCATactivity wasdetected in40-day-old callus cultures andingeneticin (G418) selected callus tissues whichalso received achimeric neomycin phosphotransferase IIgene, indicating the presence ofstable transformants. Acontrol chimeric genewith aninverted 35Spromoter failed toproduce anyCATactivity inthis system.
39 citations
TL;DR: Protoplasts isolated enzymatically from cotyledons of wild Glycine species were cultured in agarose-solidified droplets of Kao protoplast culture medium, and incubation of droplets in liquid Kao medium mixed with a B5-based formulation resulted in protiplast-derived colonies.
39 citations
TL;DR: An enzyme from Trichoderma harzianum dissolved the cell walls of a wide range of filamentous fungi and so could be used to make protoplasts and regenerated at about 1.8 times the rate of those produced by the original enzyme.
Abstract: An enzyme from Trichoderma harzianum dissolved the cell walls of a wide range of filamentous fungi belonging to Basidiomycotina, Ascomycotina, Deuteromycotina, and Zygomycotina and so could be used to make protoplasts. A lyophilized preparation of the Trichoderma enzyme had about 0.3 units/mg β-1,3-glucanase activity and 0.36 units/mg chitinase activity. About twice as many protoplasts were produced from different species of fungi by a single treatment with this enzyme than with combined commercial enzymes. The greatest number of protoplasts could be produced from most of the fungi by incubation for about 2 h t 30°C, but the number was decreased by incubation for more than 4 h or by use of a higher dose of the enzyme. An enzyme prepared by bentonite treatment from the original Trichoderma enzyme had less proteinase activity and protoplasts were fairly stable with this product during incubation for 8 h. Protoplasts produced by the proteinase-reduced preparation of the Trichoderma enzyme from three fungi regenerated at about 1.8 times the rate of those produced by the original enzyme.
39 citations
TL;DR: Protoplasts were isolated from seedling hypocotyls of Glycine canescens F.J. Herm to form calli which were tested for morphogenic potential on a range of media and produced shoots with a low but reproducible frequency.
Abstract: Protoplasts were isolated from seedling hypocotyls of Glycine canescens F.J. Herm. and cultured in agarose to form multicellular colonies. Colonies transferred to agar medium enlarged to form calli which were tested for morphogenic potential on a range of media. Protoplast-derived calli produced shoots with a low but reproducible frequency; these shoots were rooted and transferred to the greenhouse.
39 citations
TL;DR: A method was developed for the transformation of the ethanol-producing thermophilic bacteriumClostridiumthermohydrosulfuricum DSM 568 without protoplast formation and will be useful for the development of genetic exchange systems in thermophobic clostridia of biotechnological interest.
Abstract: A method was developed for the transformation of the ethanol-producing thermophilic bacteriumClostridiumthermohydrosulfuricum DSM 568 without protoplast formation. Competence for DNA uptake was induced by treatment ofCl.thermohydrosulfuricum cells with 50 mM Tris-HCl pH 8. 3. In the presence of polyethylene glycol (PEG 6000) the Tris-treated cells were transformed with the antibiotic resistance plasmids pUB110 (KmR) and pGS13 (KmR CmR) at frequencies of 4×10−6 per viable cell. This transformation method will be useful for the development of genetic exchange systems in thermophilic clostridia of biotechnological interest.
39 citations