Hey there Kristina Beranova! Dealing with Mycoplasma in egg yolk samples, huh? Well, my friend Kristina Beranova, you're diving into a niche but crucial realm. Now, I must say, extracting DNA from egg yolk can be a bit tricky, but let's cut to the chase.

So far this year, I had encountered 4 companies working on PCR testing and none of them is open to share the exact procedure for analysis. I am using some of my notes I prepared usig information available in public domain.

Firstly, there isn't a one-size-fits-all protocol for this scenario, but here's a basic rundown:

1. **Sample Preparation:**

- Separate egg yolk from the white.

- Homogenize the yolk to ensure an even distribution of the pathogen.

2. **DNA Extraction:**

- Use a reliable DNA extraction kit or a method like phenol-chloroform extraction.

- Follow the manufacturer's instructions or an established protocol.

3. **PCR Setup:**

- Design PCR primers specific to the Mycoplasma you're targeting.

- Prepare your PCR mix, including DNA template, primers, and a suitable polymerase.

4. **PCR Cycling:**

- Run your PCR program with denaturation, annealing, and extension steps.

5. **Analysis:**

- Analyze your PCR products using gel electrophoresis or another suitable method.

Remember, this is a broad overview, and specifics depend on your Mycoplasma species and the resources at your disposal. Also, the internet is a treasure trove of protocols, but the reliability can vary, so tread carefully.

Now, I'm curious—why the interest in Mycoplasma from egg yolks? Cooking up something interesting in the lab, or just a fascination with the microscopic world?

You can use alkaline NaOH method or phenol chloroform method, followed by PCR optimizing reaction

While there might not be a specific protocol for extracting mycoplasma DNA from yolk, you can adapt existing DNA extraction protocols to isolate DNA from yolk samples. Mycoplasma detection in biological samples, including eggs or yolks, often involves PCR-based methods. Here's a general guideline for mycoplasma DNA extraction and subsequent testing by PCR:

Mycoplasma DNA Extraction from Yolk:

Materials:

• Yolk samples
• DNA extraction kit (commercial kits are available)
• Microcentrifuge tubes
• Centrifuge
• Pipettes and tips
• RNase-free water

Procedure:

Mycoplasma PCR Detection:

Materials:

• Mycoplasma-specific primers
• DNA polymerase suitable for PCR
• PCR buffer and dNTPs
• DNA template (extracted mycoplasma DNA)
• PCR tubes/strips
• Thermal cycler

Procedure:

Note:

• Use precautions to avoid contamination during DNA extraction and PCR setup.
• Choose appropriate negative controls to monitor for contamination.
• Ensure that the primers used are specific to mycoplasma and do not amplify non-target DNA.

Always refer to the specific instructions provided with the DNA extraction kit and PCR reagents for detailed protocols and guidelines. If you have access to a molecular biology or microbiology laboratory, the technicians or researchers there may provide valuable assistance in optimizing the protocol for your specific sample type.

hi Kristina Beranova

Always refer to the specific

instructions provided with the DNA extraction kit and PCR reagents for detailed protocols and guidelines.

Dear colleagues Nahla Hammok Aarya Chitransh Mohamed Khedr Kaushik Shandilya , thank you very much for your recommendations and often valuable personal experience. If you have the opportunity, try the extraction kit from AnIcon, Germany. They are the only ones who have confirmed to me that it might be suitable for processing the yolk sample. I have no experience with it yet.