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Short answer
A protoplast is a cell whose cell wall has been removed, leaving the plasma membrane and all other cellular components intact. The word is used most often for plant cells, and the same idea applies to fungal and bacterial cells. Because the wall no longer holds the cell in shape, a protoplast has to be kept in a solution that balances its internal osmotic pressure.
Definition and scope: protoplast meaning
The definition of protoplast used in current plant-biology papers is short. Li et al. (2021) write that “protoplasts are plant cells that lack the cell wall, but possess plasma membrane and all other cellular components.” Sandgrind et al. (2021) put it more bluntly: “protoplasts are naked cells without cell wall.”
Three points follow from that definition.
- The wall is the only thing missing. A protoplast is not just cytoplasm or an isolated organelle. The membrane, nucleus and organelles are still there, which is why protoplasts can be transfected or fused.
- The term is not limited to plants. The wall can be removed by mechanical, chemical or enzymatic means, and the result in bacteria and fungi is also described as a protoplast (Wikipedia, Protoplast).
- Protoplasts are fragile. Without a wall they must be kept in an isotonic solution or the membrane can rupture. Li et al. (2021) report using mannitol to maintain osmotic pressure in their plant protoplast work.
The word itself is old. Wikipedia credits Hanstein with coining the term in 1880 to refer to the entire cell excluding the cell wall.
How protoplasts are made in practice
In a laboratory, protoplasts are normally produced by digesting the wall with enzymes while the cells sit in an osmotically balanced solution. The enzymes depend on what the wall is made of, so the enzyme class is the first thing that changes between organisms (see the table below). Bitesize Bio, a practical methods site, advises starting from young plant tissue or actively dividing (logarithmic-phase) cultures.
One worked example from the literature: the genome-editing study by Sandgrind et al. isolated field cress leaf protoplasts using a cellulase (Onozuka R-10) together with a macerating enzyme preparation (Macerozyme R-10), with mannitol in the plasmolysis and enzyme solutions. These are one study’s conditions, given only as an example of the reagent classes involved, not as a protocol to follow. Species, tissue and age all change what works, and the original paper’s methods section is the place to look.
Protoplasts, spheroplasts and related terms
“Protoplast” and “spheroplast” are often met together. Wikipedia distinguishes them: protoplasts have had the cell wall completely removed, while spheroplasts retain part of it. Bitesize Bio uses “spheroplast” for the spherical, wall-deficient cells it describes for bacteria and yeast. The table gives the wall-digesting enzyme class for each cell type.
| Cell type | Wall-digesting enzyme class | Source |
|---|---|---|
| Plant cells | Cellulase, pectinase, xylanase | Wikipedia, “Protoplast”; Bitesize Bio |
| Fungal cells, including yeast | Chitinase | Wikipedia, “Protoplast”; Bitesize Bio |
| Gram-positive bacteria | Lysozyme (Bitesize Bio pairs it with EDTA for bacteria) | Wikipedia, “Protoplast”; Bitesize Bio |
Because usage varies between sources, treat the two words as neighbouring terms rather than interchangeable ones, and check which one a given paper uses.
Where protoplasts matter for labs and supply decisions
Transient gene editing without regeneration
Both 2021 papers describe protoplast transient transfection as a way to deliver CRISPR components. Li et al. call it “a promising approach for delivering CRISPR complexes”. Sandgrind et al. describe using a protoplast transfection system to generate transgene-free mutants, with PEG-mediated transfection among the methods covered.
The regeneration limit
The same two papers agree on the main constraint. Protoplast regeneration “remains challenging for many plant species” (Sandgrind et al.), and Li et al. state that regeneration frequency is often species- and genotype-dependent, naming low regenerative capacity as the major obstacle in rapeseed. This splits protoplast work into two kinds with different feasibility.
| Goal | Needs regeneration into a whole plant? | What the papers report |
|---|---|---|
| Transient expression or transfection in cells | No | Described as a promising delivery route for CRISPR complexes (Li et al.) |
| Recovering edited whole plants | Yes | Regeneration is a bottleneck, often species- and genotype-dependent (Li et al.; Sandgrind et al.) |
Reagent categories a protoplast workflow draws on
A protoplast workflow in the two plant studies draws on these categories of material. This is a list of categories, not a procurement recommendation.
- Wall-digesting enzymes matched to the organism (the table above).
- An osmotic stabiliser, such as the mannitol used in both plant studies.
- Polyethylene glycol (PEG) where PEG-mediated transfection is used.
- Routine sterile consumables for handling cells, covered in the CASRAI guides to aseptic technique, conical centrifuge tubes and Petri dishes.
Common misunderstandings
- “A protoplast is just the cell contents.” The plasma membrane and all other components are present; only the wall is gone.
- “A protoplast can grow into a plant.” Sometimes, but regeneration is species- and genotype-dependent and a bottleneck for many plants.
- “Any isotonic buffer will do.” The cells must be in an osmotically balanced solution, and the plant studies cited here use mannitol. The right choice for a given species comes from the relevant paper.
- “Protoplast and spheroplast are the same word.” They are related; Wikipedia separates them by whether the wall is completely removed. Read the definition each paper gives.
FAQ
What is the meaning of protoplast?
A protoplast is a cell that has lost its cell wall but keeps its plasma membrane and other cell components. Most definitions refer to plant cells.
What is the definition of protoplast in plant biology?
Plant cells that lack the cell wall but possess the plasma membrane and all other cellular components (Li et al., 2021).
Are protoplasts only found in plants?
No. Wall removal also yields protoplasts from fungi and Gram-positive bacteria, using chitinase and lysozyme respectively, according to Wikipedia and Bitesize Bio.
Why are protoplasts used in genome editing?
Without a wall, CRISPR components can be delivered into the cells directly, for example by PEG-mediated transfection. The limit is regeneration into whole plants. For related methods see the guide to transfection methods, controls and biosafety and the electroporator overview.
For wider context on handling cells in the lab, see cell culture basics.
Sources
- Sandgrind S, et al. (2021). Frontiers in Genome Editing. PMC8635052, doi 10.3389/fgeed.2021.757540. Checked 2026-10-29.
- Li J, et al. (2021). Frontiers in Plant Science. PMC8294089, doi 10.3389/fpls.2021.680859. Checked 2026-10-29.
- Wikipedia, Protoplast. Checked 2026-10-29. Cited for the term’s origin, the protoplast and spheroplast distinction and enzyme classes by organism.
- Bitesize Bio, Breaking the wall: how to make protoplasts. Checked 2026-10-29.
