How to Prepare Plant Hormone Solutions for Tissue Culture

Introduction

Plant growth regulators synthesized by the plants are necessary for their growth and development. But, to induce the same response inside a lab in artificial conditions, it requires the addition of exogenous plant growth regulators to the media.

Today, several natural and synthetic plant growth regulators (PGRs), plant hormones, or phytohormones are available for use in tissue culture experiments. But how to prepare a PGRs solution is a most common problem or confusion faced by our culturists!

So, in this article, we’ve covered how to prepare the stock solution of plant growth regulators and the stock solution of NaOH (solvent) required to dissolve the powdered form hormone.

How to Prepare a Stock Solution of PGRs?

To prepare the stock solution of any plant growth regulators follows the steps given below:

• Take 50 ml of falcon tube for each hormone for which you are willing to prepare the stock.
• Label each tube properly with the name of the hormone.
• To prepare 1mg/L plant growth hormone working solution, prepare 1g/L of solution with water.
• In the video, we are measuring 0.05 gram (50 mg) of powdered form of the hormone to prepare a 50 ml stock solution.
• You can use a cup and spatula to measure each plant growth regulator. But, you should change the cup and clean or change the spatula every time you measure another plant growth regulator to avoid any cross-contamination of your stock solutions.

Note: From a PCT’s 100-gram bottle of plant growth regulators, you can prepare 100 liters of stock solution. However, you only need 1 ml of stock solution to make 1 liter of media. So, overall you can prepare 100K liters of media per bottle.

• The next step is dissolving the powder form of the growth regulator. It won’t directly dissolve in water, so you need a solvent.
• Use NaOH as a solvent to dissolve the powder form of the growth regulator.
• Using a micropipette measure 1ml of NaOH and add it to the 50 ml falcon tube containing the powdered growth regulator. Mix it properly and change pipette tips before moving to mix the next growth regulators to avoid any cross-contamination between hormones (if you are preparing a plant hormone stock solution of more than one hormone).

You can also use the small stirrer or shake the tube or bottle to dissolve the powdered hormone into the solvent.

Note: The procedure to prepare 1M NaOH is given below. So, take 1mlNaOH from the prepared 1M solution to dissolve plant growth regulators.

• Then add 49 ml (because you’ve already added 1 ml of NaOH to dissolve the powdered hormone) of distilled water using a pipette to make 50 ml of the stock solution of the hormone.
• Then, it’s time to sterilize the stock solution as they are not sterile. You should know about your hormone if it’s heat resistant or not.
• If the hormone is not heat resistant then, you need to syringe filter the solution after the media has cooled down to at least 60 degrees celsius and if it’s heat resistant then you can autoclave the solution.
• Then your stock solution is ready to use for your experiment. You can dilute it to bring it to your desired concentration.

How to Prepare 1M NaOH solution?

Sodium hydroxide (NaOH) is used to increase the pH of the culture media or dissolve the powdered form of growth regulator.

• To prepare the NaOH stock solution you need NaOH pellets, a spatula, Erlenmeyer flask.
• Add 100 ml distilled water to the Erlenmeyer flask using the pipette.
• To make one molar solution of NaOH you need the formula:

Molarity = no. of moles of solute / 1 liter

And, one mole of sodium hydroxide = 40 grams

So, if you want to prepare 1M of NaOH solution, you need 40 grams of NaOH dissolved in 1L of the solution.

But, here we are preparing only 100 ml of the stock solution, so for that, you only need 4 grams of NaOH dissolved in 100 ml water.

• Measure 4 grams of NaOH using a spatula on a weighing machine.
• Add the measured NaOH pellets into water and using a stirred make it a homogenous solution.
• After the pellets are dissolved, transfer the solution to the storage bottle.
• Your 1 molar NaOH solution is ready to use for your experiments.

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