Arabidopsis Pollen Germination Medium
Introduction
There are a handful of Pollen Germination Media (PGM) formulated for one of the widely used model plant, Arabidopsis. In this article, we have provided the composition of one of the robust PGM. This PGM composition is adopted from a publication by Leonor 2007 (see the reference at the end of the article). If you are a beginner, I highly recommend you to read the full article since it shows the effect of temperature, light, pH and other factors on pollen germination and pollen tube growth. We have placed reagents required for preparation in a tabular form. This also contains information on Molarity of each reagent and their quantities to be added for making 10 mL of stock solution. We have also provided information about different dilutions that can be made from the stock.
Composition
| Reagent | M.W | Working concentration | Stock solution | Add for 10 ml stock | Add from stock for 1X of 50ml |
|---|---|---|---|---|---|
| Sucrose | 342.3 | 10% | - | - | 5 g |
| H2BO3 | 61.8 | 0.01% | 1% | 0.1 g | 0.5 ml |
| CaCl2(anhydrous) | 110.98 | 5mM | 1M | 1.1 g | 250 µl |
| KCl | 74.5 | 5mM | 1M | 0.745 g | 250 µl |
| MgSO4 .7H2O | 246.47 | 1mM | 1M | 2.465 g | 50 µl |
| Noble Agar | - | 1.50% | - | - | 0.75 g |
Table 1. Composition of Arabidopdsis pollen germination medium.
pH
The pH of the medium should be adjusted to 7.5 using dilute HCl and KOH.
Preparation
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 measuring 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 a magnetic stirrer or vortex mixer.
- Adjust pH to 7.5 using HCl and KOH (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.
Storage
Store PGM (Pollen Germination Medium) stock solutions at 4˚C for 1-2 weeks.