Protocols & Procedures:
The procedures described below are generic. Slight modifications might be needed for your specific plant species. For assistance, contact the PCT team at info@plantcelltechnology.com
PPM™ significantly simplifies the tissue culture working procedures as follows:
1. Media containing PPM™ may be dispensed outside the laminar flow hood (LFH) exposed to the ambient air. The plates should be covered soon after agar solidification. In the event that media dispensing is done by a pump, we recommend passing autoclaved hot water through the hoses prior to and after media dispensing.
2. Heat sensitive or heat stable liquid media containing PPM™ does not need to be filter sterilized or autoclaved provided that it will be stored in sterile containers and that the stock solutions are not contaminated. In rich media containing 200 mg/liter or more of amino acids or proteins, it is recommended to filter the media with the PPM™.
3. If working in the LFH the utensils (forceps or scalpels) do not need to be flamed. They should be periodically dipped in 70% alcohol. The LFH does not need to be certified and the work can be done as well outside the LFH on a clean surface for a period not exceeding 1 hour.
4. PPM™ is less effective when exposed to high density of bacteria or fungi spores found regularly on seed's coat. For in vitro germination, seeds should be conventionally surface sterilized with EPA registered bleach. Therefore, in the presence of PPM™ (in the germination medium), the seeds can be rinsed under tap water in a non-sterile strainer and left to dry preferably in the LFH. If the utensil ends have touched active bacteria, fungi culture or otherwise suspected of being contaminated, they should be sterilized by autoclave or by use of an electric heating element.
5. General Dosage levels: With the exception of endogenous contamination, the recommended dose range is 0.05%-0.2%. (For callus proliferation, organogenesis and embryogenesis, the recommended range is 0.05-0.075%.) To eliminate higher endogenous contamination densities, higher doses of PPM are needed (see paragraph 6 below).
Note
Paragraphs 6(b) through 10 below are intended for ornamental plants only.
Refer to notes 2 and 3 below.
7. To eliminate Agrobacterium:
To obtain better results after cultivation with Agrobacterium, add 0.2% PPM v/v directly from the bottle to an antibiotic cocktail.
General Notes:
For the first transfer following the sterilization with PPM™, we recommend to insert the explants entirely into a semi-solid medium.
Summary
PPM™ most definitely will facilitate the work in any plant tissue laboratory and should significantly increase technician and laboratory productivity. However, conditions in each lab may vary which could have a bearing on the effectiveness of PPM™. It is advisable that staff follow the above guidelines initially and adjust parameters accordingly.
When used as recommended:
- PPM™ is effective against airborne contamination, waterborne contamination and contamination introduced from human contact.
- At recommended doses (0.5 - 2ml/l ), PPM™ does not impair in vitro seed germination, callus proliferation, callus regeneration, and axillary or adventitious buds' induction.
Safety and Handling - See MSDS
DO NOT CONCENTRATE THE MATERIAL
DISPOSAL INFORMATION:
Dispose of media containing PPM™ in the same manner in which you dispose of media without PPM™. In an emergency, contact the following numbers: 1 (202) 621-5490 or +1.800.746.8535. A toxicological assessment has been performed by a qualified toxicologist. This assessment is available upon request. For more information on PPM™, or to request test results contact: Tel: 1.202.778.8522 ex. 0, Fax: 1.202.429.9812