Protoplast

About: Protoplast is a research topic. Over the lifetime, 5474 publications have been published within this topic receiving 122468 citations.

Isolation, culture, and transient transformation of plant protoplasts.

Abstract: Transient gene expression in protoplasts, which has been used in several plant species, is an important and versatile tool for rapid functional gene analysis, protein subcellular localization, and biochemical manipulations. This unit describes transient gene expression by electroporation of DNA into protoplasts of Arabidopsis or tobacco suspension-cultured cells and by polyethylene glycol (PEG)-mediated DNA transformation into protoplasts derived from rice leaf sheaths. PEG-mediated DNA transformation for transient gene expression in rice protoplasts in suspension culture is also described as an alternative technique. Methods for collecting intracellular and secreted proteins are also provided.

Transfer of cytoplasm from newBeta CMS sources to sugar beet by asymmetric fusion : 1. Shoot regeneration from mesophyll protoplasts and characterization of regenerated plants.

Abstract: For our program on the transfer of cytoplasmic male sterility (CMS) by cybridization inBeta vulgaris L. (sugar beet), we have developed a procedure for the isolation and culture of mesophyll protoplasts of sugar beet followed by shoot regeneration. A prerequisite proved to be the presence in the media of n-propylgallate (nPG), a lipoxygenase inhibitor. Sustained divisions were found in all accessions that were tested. Plating efficiencies and regeneration ability varied greatly from one experiment to the other and appeared to be accession-dependent. Shoots could be easily transferred to soil. A majority of the regenerants (72%) retained the diploid chromosome number. Somaclonar variation in phenotype was low (4.9%). Mitochondrial DNA probes, capable of discriminating different cytoplasms ofBeta spp. showed no rearrangements due to the protoplast and in vitro culture phase, indicating that these probes can be used to identify cybrids after asymmetric fusions. The data presented here open up possibilities for genetic engineering using protoplasts in one of the world's most important arable crops.

Amino acid auxotrophs from protoplast cultures of Nicotiana plumbaginifolia , Viviani

Abstract: Several amino acid requiring auxotrophs have been isolated from unsupplemented protoplast cultures of haploid Nicotiana plumbaginifolia following incubation with BUdR (1-5x10-5, 2 days) and recovery on complete medium. The auxotrophic lines required the following amino acid(s) for growth: his, ile, leu, ile+val, met or try. Met− is a new type isolated in higher plants. The same absolute amino acid requirement was observed in plants regenerated from auxotrophic cultures. Precursor feeding tests, enzyme assays, and/or metabolic complementation through protoplast fusion were used to identify the genetic lesion leading to auxotrophy. Mutant seeds were obtained from supplemented Met− plants. Seeds were also collected from selfed plants regenerated from various complementing fusion products, and a His− revertant. Genetic analysis indicated that under natural conditions of seed formation amino acid auxotrophy-in contrast to NR deficiency-failed to segregate in progeny tests.