Arabidopsis Pollen Germination Medium (Li H 1999)
The pollen germination medium (PGM) is used for allowing the pollen to germinate either by in vitro or semi-in vitro methods. This web page deals with the composition and preparation of PGM as mentioned in the article titled “Control of Pollen Tube Tip Growth by a Rop GTPase–Dependent Pathway That Leads to Tip-Localized Calcium Influx” published in the plant cell by Li H1999 et al.
For your convenience, we have given the molecular weights of each reagent and its most used hydrated forms. It is up to the user to determine whether hydrated forms make any difference for the experiments or not.
The pH of the medium should be adjusted to 7.0 using dilute HCl.
|Reagent||M.W||Working concentration||Stock solution||Add for 10 ml stock||Add from stock for 1X of 50ml|
|H2BO3||61.8||0.01%||1%||0.1 g||0.5 ml|
|CaCl2(anhydrous)||110.98||1mM||1M||1.1 g||50 µl|
|CaCl2 .2H2O||147.01||1 mM||1M||1.47 g||50 µl|
|Ca(NO3)2||164.08||1mM||1M||1.64 g||50 µl|
|Ca(NO3)2 .4H2O||236.15||1mM||1M||2.36 g||50 µl|
|MgSO4||120.36||1mM||1M||1.20 g||50 µl|
|MgSO4 .7H2O||246.47||1mM||1M||2.46 g||50 µl|
|Noble Agar||-||0.50%||-||-||0.25 g|
- Boric acid
- Calcium chloride
- Calcium nitrate
- Magnesium sulphate
- Noble agar
- Distilled water
Instruments and other requirements
- Weighing balance. paper boats.
- Graduated cylinder and a glass beaker.
- pH meter. Magnetic stirrer and magnetic bead/pellet.
- Tissue paper.
Use ddH2O only for making PGM
You can choose to make 1X (working concentration) or make stocks and use them to make 1X whenever you want. Making stock solutions eliminate the possibility of errors that may happen while weighing very low quantities of reagents. Making working concentration from stocks also helps you to add some other reagents without further diluting working concentration.
- Determine whether you want to start with stock solutions or directly making working concentration.
- Weigh the reagents accordingly (Refer to the table above).
- Dissolve salts by using magnetic stirrer or vortex mixer.
- Adjust pH to 7.0 using HCl and KOH/NaOH (pH is one of the critical factors)
- Make the volume as planned.
- Sterilization of media is not required for immediate use. However, doing filter sterilization might increase its shelf life by preventing microbial growth.
- Aliquot to smaller volumes if needed (frequent opening and closing in open air might result in microbial growth).
- Noble agar is preferable over agar as a solidifying agent.
Store PGM (Pollen Germination Medium) stock solutions at 4˚C for 1-2 weeks.
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