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.  

pH

The pH of the medium should be adjusted to 7.0 using dilute HCl.

Composition

Arabidopsis Pollen Growth Medium (PGM) composition and preparation (as per Li H 1999 et al.)
pH is recommended to be 7.0. achieve this pH using HCl and KOH.
Arabidopsis Pollen Growth Medium (PGM) composition and preparation (as per Li H 1999 et al.)
pH is recommended to be 7.0. achieve this pH using HCl and KOH.
Reagent M.W Working concentration Stock solution Add for 10 ml stock Add from stock for 1X of 50ml
Sucrose 342.3 18% - - 9 g
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

Reagents

  • Sucrose
  • Boric acid
  • Calcium chloride
  • Calcium nitrate
  • Magnesium sulphate
  • Noble agar
  • Distilled water
  • HCl

Instruments and other requirements

  • Weighing balance. paper boats.
  • Graduated cylinder and a glass beaker.
  • pH meter. Magnetic stirrer and magnetic bead/pellet.
  • pipette.
  • Tissue paper.

Procedure

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.  

  1. Determine whether you want to start with stock solutions or directly making working concentration.
  2. Weigh the reagents accordingly (Refer to the table above).
  3. Dissolve salts by using magnetic stirrer or vortex mixer.
  4. Adjust pH to 7.0 using HCl and KOH/NaOH (pH is one of the critical factors)
  5. Make the volume as planned.
  6. Sterilization of media is not required for immediate use. However, doing filter sterilization might increase its shelf life by preventing microbial growth.
  7. Aliquot to smaller volumes if needed (frequent opening and closing in open air might result in microbial growth).
  8. Noble agar is preferable over agar as a solidifying agent.

Storage

Store PGM (Pollen Germination Medium) stock solutions at 4˚C for 1-2 weeks.

References

Control of Pollen Tube Tip Growth by a Rop GTPase–Dependent Pathway That Leads to Tip-Localized Calcium Influx

site icon

Share this page on

Leave a Comment