PPM ™ is a thermoset preservative/biocide that can be used to effectively prevent or reduce microbial contamination in plant tissue cultures. In optimal doses, PPM, which stands for Plant Preservative Mixture ™, is an extremely effective preservative/biocide that does not harm in vitro seed germination, callus proliferation, and regeneration.
Despite the strictest use of sterile techniques, contamination of plant cells and plant tissue cultures remains a persistent problem that can result in losses ranging from a small number of cultures to complete batches. PPM ™ can and should be used as a standard ingredient in plant tissue culture media. It is substantially less expensive and potentially less dangerous than commonly used antibiotics. PPM ™ was designed primarily to inhibit air, water, and human contact-induced pollution. However, in many cases, it can also be used to reduce endogenous contamination.
Mechanism of action
PPM ™ is a broad-spectrum preservative and biocide, which kills bacteria and fungi cells, prevents spore germination, and in higher concentrations, it can eliminate endogenous contamination of explants. It is heat stable and therefore can be autoclaved.
Previous research has shown that the active ingredients in PPM ™ penetrate the cells of fungi or bacteria and inhibit key enzymes in central metabolic cycles such as the citric acid cycle and the electron transport chain. Our data indicate that PPM can also inhibit the transport of monosaccharides and amino acids from the medium in fungal or bacterial cells. As with any biocide, a critical ratio of PPM ™ molecules per microbial cell is required. to eliminate bacteria and fungi.
Note that a given volume of PPM ™ doses has a constant number of PPM ™ molecules, while the number of spores introduced into tissue culture through endogenous contamination is highly varied. Therefore, explants must not be “squeezed” into a beaker. There must be enough volume to move freely of the solution around the explanted material.
Advantages of PPM over antibiotics
- PPM ™ is broad-based and effective against fungi.
- PPM ™ is less expensive than antibiotics, making it affordable for routine and extensive use.
- Since PPM ™ targets and inhibits multiple enzymes, the formation of resistant mutants towards PPM ™ are highly unlikely.
- PPM ™ is heat stable and can generally be autoclaved with media.
- Media containing PPM ™ can be dispensed outside of laminar flow hoods (LFH) under ambient conditions [however, the use of orchid hoods is recommended laboratories]. Culture plates and containers should be covered shortly after the agar solidification. When dispensing media with pumps, PCT recommends passing Autoclave hot water through the hoses before and after dispensing the medium.
- Heat-sensitive or thermoset liquid media containing PPM ™ need not be sterilized by passing through Millipore filters or autoclaved provided that it should be stored in sterile containers and that the stock solutions are not contaminated. In rich media containing 200 mg l – 1 or more of amino acids or proteins. It is recommended to filter and sterilize the medium with PPM ™ [as few orchids the media is so rich that this should not be necessary].
- When working on LFH, there is no need to flare tools (forceps or scalpels) [to use with orchids, flaming is recommended]. They should be periodically immersed in 70% alcohol. The LFH does not need to be certified [LFH used with orchids do does not require certification anyway]. The work can be performed outside of the LFH on a clean surface for a period not exceeding 1 h.
- PPM ™ is an acidic liquid solution (pH 3.8) that should be stored at 4 ° C (see safety information below). At the recommended dose of 0.05 to 0.2% (v / v), PPM ™ added to the medium before or after autoclaving to avoid air and endogenous contamination at low inoculum densities.
PPM ™ will definitely make work easier in any plant tissue laboratory and should significantly increase the productivity of technicians and laboratories. However, the conditions in each laboratory may vary and this could influence the effectiveness of PPM. It is recommended that personnel follow the above guidelines to maximize the effectiveness of PPM ™ for a specific laboratory and particular applications. Test results show that:
- PPM ™ is effective against air and water pollution and contaminants introduced by human contact.
- If used correctly, PPM ™ will release seeds or explants of endogenous substances contamination.
- At the recommended doses (0.5–2 ml l – 1), PPM ™ does not damage the seed in vitro germination, callus proliferation, callus regeneration or axillary induction or adventitious buds.
- The formulation of certain concentrations of PPM and their use in tissue culture to preventing or eliminating microbial contamination is protected by US Patent No. 5,750,402. It is also patent pending in many countries around the world.