Protoplast (), is a

biological Biology is the scientific study of life. It is a natural science with a broad scope but has several unifying themes that tie it together as a single, coherent field. For instance, all organisms are made up of cells that process hereditary in ...

term coined by Hanstein in 1880 to refer to the entire cell, excluding the cell wall. Protoplasts can be generated by stripping the

cell wall A cell wall is a structural layer surrounding some types of cells, just outside the cell membrane. It can be tough, flexible, and sometimes rigid. It provides the cell with both structural support and protection, and also acts as a filtering mech ...

from

plant Plants are predominantly photosynthetic eukaryotes of the kingdom Plantae. Historically, the plant kingdom encompassed all living things that were not animals, and included algae and fungi; however, all current definitions of Plantae exclud ...

bacterial Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one biological cell. They constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria were among ...

, or

fungal A fungus ( : fungi or funguses) is any member of the group of eukaryotic organisms that includes microorganisms such as yeasts and molds, as well as the more familiar mushrooms. These organisms are classified as a kingdom, separately from th ...

cells by mechanical, chemical or enzymatic means. Protoplasts differ from

spheroplasts A spheroplast (or sphaeroplast in British usage) is a microbial cell from which the cell wall has been almost completely removed, as by the action of penicillin or lysozyme. According to some definitions, the term is used to describe Gram-negative ...

in that their cell wall has been completely removed. Spheroplasts retain part of their cell wall. In the case of

Gram-negative bacteria Gram-negative bacteria are bacteria that do not retain the crystal violet stain used in the Gram staining method of bacterial differentiation. They are characterized by their cell envelopes, which are composed of a thin peptidoglycan cell wall ...

l spheroplasts, for example, the

peptidoglycan Peptidoglycan or murein is a unique large macromolecule, a polysaccharide, consisting of sugars and amino acids that forms a mesh-like peptidoglycan layer outside the plasma membrane, the rigid cell wall (murein sacculus) characteristic of most ...

component of the cell wall has been removed but the outer membrane component has not.

Enzymes for the preparation of protoplasts

Cell walls are made of a variety of

polysaccharide Polysaccharides (), or polycarbohydrates, are the most abundant carbohydrates found in food. They are long chain polymeric carbohydrates composed of monosaccharide units bound together by glycosidic linkages. This carbohydrate can react with wa ...

s. Protoplasts can be made by degrading cell walls with a mixture of the appropriate polysaccharide-degrading

enzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. A ...

s: During and subsequent to digestion of the cell wall, the protoplast becomes very sensitive to

osmotic Osmosis (, ) is the spontaneous net movement or diffusion of solvent molecules through a selectively-permeable membrane from a region of high water potential (region of lower solute concentration) to a region of low water potential (region of ...

stress. This means cell wall digestion and protoplast storage must be done in an isotonic solution to prevent rupture of the

plasma membrane The cell membrane (also known as the plasma membrane (PM) or cytoplasmic membrane, and historically referred to as the plasmalemma) is a biological membrane that separates and protects the interior of all cells from the outside environment (t ...

Uses for protoplasts

Protoplasts can be used to study

membrane A membrane is a selective barrier; it allows some things to pass through but stops others. Such things may be molecules, ions, or other small particles. Membranes can be generally classified into synthetic membranes and biological membranes. B ...

biology, including the uptake of

macromolecule A macromolecule is a very large molecule important to biophysical processes, such as a protein or nucleic acid. It is composed of thousands of covalently bonded atoms. Many macromolecules are polymers of smaller molecules called monomers. The ...

s and

virus A virus is a submicroscopic infectious agent that replicates only inside the living cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, including bacteria and archaea. Since Dmitri Ivanovsky's 1 ...

es . These are also used in

somaclonal variation Somaclonal variation is the variation seen in plants that have been produced by plant tissue culture. Chromosomal rearrangements are an important source of this variation. The term somaclonal variation is a phenomenon of broad taxonomic occurrenc ...

. Protoplasts are widely used for DNA

transformation Transformation may refer to: Science and mathematics In biology and medicine * Metamorphosis, the biological process of changing physical form after birth or hatching * Malignant transformation, the process of cells becoming cancerous * Trans ...

(for making

genetically modified organism A genetically modified organism (GMO) is any organism whose genetic material has been altered using genetic engineering techniques. The exact definition of a genetically modified organism and what constitutes genetic engineering varies, with ...

s), since the cell wall would otherwise block the passage of DNA into the cell. In the case of plant cells, protoplasts may be regenerated into whole plants first by growing into a group of plant cells that develops into a

callus A callus is an area of thickened and sometimes hardened skin that forms as a response to repeated friction, pressure, or other irritation. Since repeated contact is required, calluses are most often found on the feet and hands, but they may o ...

and then by regeneration of shoots ( caulogenesis) from the callus using

plant tissue culture Plant tissue culture is a collection of techniques used to maintain or grow plant cells, tissues or organs under sterile conditions on a nutrient culture medium of known composition. It is widely used to produce clones of a plant in a method known ...

methods. Growth of protoplasts into callus and regeneration of shoots requires the proper balance of plant growth regulators in the tissue culture medium that must be customized for each species of plant. Unlike protoplasts from

vascular plant Vascular plants (), also called tracheophytes () or collectively Tracheophyta (), form a large group of land plants ( accepted known species) that have lignified tissues (the xylem) for conducting water and minerals throughout the plant. They al ...

s, protoplasts from

moss Mosses are small, non-vascular flowerless plants in the taxonomic division Bryophyta (, ) '' sensu stricto''. Bryophyta (''sensu lato'', Schimp. 1879) may also refer to the parent group bryophytes, which comprise liverworts, mosses, and hor ...

es, such as ''

Physcomitrella patens ''Physcomitrium patens'', (synonym: ''Physcomitrella patens'' ) the spreading earthmoss, is a moss (bryophyte) used as a model organism for studies on plant evolution, development, and physiology. Distribution and ecology ''Physcomitrella pat ...

'', do not need

phytohormone Plant hormone (or phytohormones) are signal molecules, produced within plants, that occur in extremely low concentrations. Plant hormones control all aspects of plant growth and development, from embryogenesis, the regulation of organ size, pat ...

s for regeneration, nor do they form a callus during

regeneration Regeneration may refer to: Science and technology * Regeneration (biology), the ability to recreate lost or damaged cells, tissues, organs and limbs * Regeneration (ecology), the ability of ecosystems to regenerate biomass, using photosynthesis ...

. Instead, they regenerate directly into the filamentous

protonema A protonema (plural: protonemata) is a thread-like chain of cells that forms the earliest stage of development of the gametophyte (the haploid phase) in the life cycle of mosses. When a moss first grows from a spore, it starts as a ''germ tube'', ...

, mimicking a germinating moss spore. Protoplasts may also be used for

plant breeding Plant breeding is the science of changing the traits of plants in order to produce desired characteristics. It has been used to improve the quality of nutrition in products for humans and animals. The goals of plant breeding are to produce cro ...

, using a technique called

protoplast fusion Somatic fusion, also called protoplast fusion, is a type of genetic modification in plants by which two distinct species of plants are fused together to form a new Hybrid (biology), hybrid plant with the characteristics of both, a somatic hybri ...

. Protoplasts from different species are induced to fuse by using an

electric field An electric field (sometimes E-field) is the physical field that surrounds electrically charged particles and exerts force on all other charged particles in the field, either attracting or repelling them. It also refers to the physical field fo ...

or a solution of

polyethylene glycol Polyethylene glycol (PEG; ) is a polyether compound derived from petroleum with many applications, from industrial manufacturing to medicine. PEG is also known as polyethylene oxide (PEO) or polyoxyethylene (POE), depending on its molecular we ...

.Hain R, Czernilofsky AP, et al. (1985). "Uptake, integration, expression and genetic transmission of a selectable chimaeric gene by plant protoplasts". ''Molecular and General Genetics'' 199:161–168. This technique may be used to generate

somatic hybrid Somatic fusion, also called protoplast fusion, is a type of genetic modification in plants by which two distinct species of plants are fused together to form a new hybrid plant with the characteristics of both, a somatic hybrid. Hybrids have be ...

s in tissue culture. Additionally, protoplasts of plants expressing fluorescent proteins in certain cells may be used for Fluorescence Activated Cell Sorting (FACS), where only cells fluorescing a selected wavelength are retained. Among other things, this technique is used to isolate specific cell types (e.g.,

guard cells Guard cells are specialized plant cells in the epidermis of leaves, stems and other organs that are used to control gas exchange. They are produced in pairs with a gap between them that forms a stomatal pore. The stomatal pores are largest when ...

from leaves,

pericycle The pericycle is a cylinder of parenchyma or sclerenchyma cells that lies just inside the endodermis and is the outer most part of the stele of plants. Although it is composed of non-vascular parenchyma cells, it's still considered part of the vas ...

cells from roots) for further investigations, such as transcriptomics.

References

{{Reflist Cell biology Membrane biology Molecular biology Plant physiology Plant reproduction

Enzymes for the preparation of protoplasts

Uses for protoplasts

See also

References