LIVE Plant Tissue Culture Biocoupler™ Liquid Culture Subculturing with Francisco

Introduction

This tutorial provides a step-by-step guide on how to perform a media change for BioCouplers™ in plant tissue culture, as demonstrated by Francisco from Plant Cell Technology. BioCouplers™ are advanced systems designed to optimize the growth conditions for plant tissues, making them essential for effective tissue culture operations. This guide will help you understand the process and advantages of using BioCouplers™ in your own tissue culture practices.

Step 1: Gather Necessary Materials

Before starting, ensure you have all required materials ready. Here’s what you need:

• Supreme-Grade Agar
• Gellan Gum
• PPM™ (Plant Preservative Mixture)
• MS Media (Murashige and Skoog medium)
• Vented Test Tubes (100 ml)
• Round Snap-Lock Containers
• BioCouplers™
• IBA Solution (Indole-3-butyric acid)
• BAP Solution (Benzylaminopurine)

Practical Tips

• Ensure all materials are sterile to avoid contamination.
• Familiarize yourself with each component and its purpose in the media.

Step 2: Prepare the Media

To change the media in your BioCouplers™, start by preparing the new culture media.

1. Measure Ingredients:

   • Follow the standard recipes for MS media, using the appropriate amounts of agar, gellan gum, and any growth regulators like IBA or BAP.

2. Mix and Heat:

   • Combine the dry ingredients in a suitable container and add distilled water.
   • Heat the mixture until the agar dissolves completely.

3. Sterilize:

   • Transfer the mixture to a suitable vessel and sterilize it using an autoclave.

Common Pitfalls to Avoid

• Ensure that the media is not overheated to avoid degradation of growth regulators.
• Always check for bubbles in the media after heating, as they can affect culture quality.

Step 3: Remove Old Media from BioCouplers™

Carefully remove the old media from the BioCouplers™ to prepare for the new media.

1. Open BioCouplers™:

   • Use aseptic techniques to open the BioCouplers™ without introducing contaminants.

2. Discard Old Media:

   • Gently pour out the old media into a waste container, ensuring that you do not disturb the plant tissues inside.

Practical Advice

• Work quickly but carefully to minimize exposure to air and potential contaminants.

Step 4: Add New Media to BioCouplers™

Once the old media is removed, it’s time to add the newly prepared media.

1. Pour New Media:

   • Slowly pour the new media into the BioCouplers™, ensuring that it covers the plant tissues adequately.

2. Seal BioCouplers™:

   • Close the BioCouplers™ securely to maintain a sterile environment.

Real-World Application

• This step is crucial for promoting healthy growth and development in plant tissues, enabling successful subculturing.

Step 5: Monitor and Adjust Conditions

After the media change, monitor the conditions to ensure optimal growth.

1. Check Environmental Factors:

   • Keep an eye on temperature, light, and humidity levels as these can significantly affect tissue culture success.

2. Adjust as Necessary:

   • Be prepared to adjust the environmental conditions based on the specific needs of the plant species you are culturing.

Key Tips

• Regularly monitor the cultures for signs of contamination or growth issues.
• Engage with the plant tissue culture community for ongoing support and tips.

Conclusion

Performing a media change in BioCouplers™ is an essential procedure for successful plant tissue culture. By following these steps, you ensure that your plant tissues receive the optimal conditions for growth. Remember to gather your materials, prepare your media carefully, and monitor the environmental conditions for the best results. For further learning, consider participating in master classes or engaging with online communities focused on plant tissue culture. Happy culturing!