To develop a gene transformation method for Flammulina velutipes, we constructed a vector with hph gene under control of the trp1 gene promoter. The vector was integrated into protoplast derived from mycelia by the calcium-polyethylene glycol method, as it has not been reported for F. velutipes. Transformation efficiency was much improved when transformation was performed by the restriction enz...

The high osmotic pressure provided for in vitro transformation processes to support naked protoplasts of Saccharomyces cerevisiae was found to hurt regeneration of the protoplasts to walled cells. Preadaptation of the cells by growth under the same hypertonic conditions as those used for transformation doubled the rate of protoplast regeneration and in turn resulted in a higher yield of transfo...

We have established that polypeptides whose synthesis is reduced by 2,4-dichlorophenoxyacetic acid during in vitro culture of tobacco mesophyll protoplasts are secreted into the vacuole where they constitute the bulk of labeled proteins. In addition, these proteins continue to be synthesized in protoplast-derived cultured cells and their synthesis is strictly correlated with the size of the cel...

Plant protoplasts show physiological perceptions and responses to hormones, metabolites, environmental cues, and pathogen-derived elicitors, similar to cell-autonomous responses in intact tissues and plants. The development of defined protoplast transient expression systems for high-throughput screening and systematic characterization of gene functions has greatly contributed to elucidating pla...

A rapid method for nuclease digestion of Aspergillus nidulans chromatin is described. It overcomes the need for nuclear purification or protoplast preparation. The method is valid for the analysis of the nucleosomal repeat length in bulk chromatin, and allows the analysis of nucleosome phasing at a specific locus.

The cytoplasm of the interior of maturing pine tracheids is revealed by scanning electron microscopy as a lamellate and filamentous intravacuolar reticulum immersed in vacuolar sap. Formation of cytoplasmic spherules and the breakdown of the cytoplasmic reticulum initiate protoplast disintegration that terminates in tracheid maturation. The process is believed to involve vacuolar membrane trans...

Plasmid pIL7 (33 kilobases) from Streptococcus lactis enhances UV resistance and prophage stability. A 5.4-kilobase pIL7 fragment carrying genes coding for both characters was cloned into S. lactis, using plasmid pHV1301 as the cloning vector. The recombinant plasmid was subsequently transferred to three other S. lactis strains by transformation or protoplast fusion. Cloned genes were expressed...

Periplasmic aminopeptidase II of yeast was isolated from protoplast supernatants by ammonium sulfate precipitation, DEAE-Sephacel chromatography and affinity chromatography on immobilized bestatin. This isolation method is more rapid than conventional procedures and, in addition, avoids contact of the enzyme with yeast proteinases. Thus the preparation obtained is likely to represent the native...