You could use a transport medium containing bacterial charcoal (e.g. the one for Campylobacter) if your sample site is far.

If the site is near then you just collect the sample (warer or soil) and process it.

I do not think a few hours can affect DNA or RNA. As long as your sample is not old, you are fine.

Or you could freeze the sample at -20C and process it later.

thank you for your kind help, my sample is located in a hot desert far from well-equipped cities.

1- Jafar Sabouri You can take Petri dishes containing the media to the area, and they must be in a container that maintains the temperature. Also, wrap the dishes containing the media in aluminum foil, which helps maintain the temperature until arriving at the laboratory.

2- You can take a sample in a suitable container to the laboratory

3- You can take a cotton swab to the site, take the swabs, and then go to the laboratory within 24 hours.

Dear Abdelmoniem,

Thank you very much for your useful answer.

I asked Chat GPT about how to use preservers of chromosomes and genes when sampling from the environment to the lab which answered as follows:

"Adding preservatives: After collecting the samples and conducting an initial examination, add a solution containing DNA and RNA preservatives to the samples. This action helps preserve the DNA and RNA structures of the microorganism samples during transportation to the laboratory.

The use of DNA and RNA preservatives is for the preservation of chromosomes and genes. These preservatives include substances such as glycerol, dimethyl sulfoxide (DMSO), polyethylene glycol (PEG), and other materials that are used in the processes of preserving and storing biological samples."

Can you guide me about these preservatives and how to prepare them?

Preserving microorganisms like bacteria, cyanobacteria, and algae from environmental samples to laboratory settings requires careful planning and execution. Here is a detailed guide on the practical methods for sample collection, packaging and transporting, preservation, and laboratory analysis:

1. Sample Collection

Bacteria

• Water Samples: Use sterile bottles or containers to collect water samples. Avoid touching the inside of the container or the cap.
• Soil Samples: Collect soil using a sterile spatula or corer. Place the soil in sterile containers or plastic bags.
• Surfaces: Use sterile swabs or wipes to collect bacteria from surfaces. Place the swabs in sterile transport media or tubes.

Cyanobacteria and Algae

• Water Samples: Use sterile bottles for water collection. For planktonic samples, use a plankton net to concentrate the organisms before transferring them to bottles.
• Surface Samples: For biofilm or periphyton samples, scrape the surface gently with a sterile tool and transfer to sterile containers.
• Sediment Samples: Use a sterile corer or spatula to collect sediment samples and place them in sterile containers.

2. Packaging and Transporting

• Packaging: Ensure that all containers are properly sealed to prevent contamination and leakage. Use insulated coolers with ice packs if the samples need to be kept cool.
• Labeling: Clearly label each sample with relevant information, including the date, time, location, and type of sample.
• Transport Conditions: Transport samples to the laboratory as soon as possible. For samples that need to be kept cool, maintain them at 4°C during transport.

3. DNA and RNA Preservation

Preservation Methods:

• For Bacteria and Algae:DNA: Use commercial DNA preservation solutions like DNA/RNA Shield, RNAlater, or similar. These solutions stabilize nucleic acids at room temperature. RNA: RNA is more labile than DNA and requires immediate stabilization. Use RNA stabilization solutions (like RNAlater) and store samples at -80°C for long-term preservation.

Immediate Processing:

• If immediate preservation solutions are not available, process samples as soon as possible in the laboratory. For DNA, freeze samples at -20°C; for RNA, freeze at -80°C.

4. Laboratory Analysis

Microscopy Study:

• Preparation: For microscopy, prepare wet mounts, stained slides, or fixed samples as needed.
• Staining: Use specific stains for better visualization (e.g., Gram stain for bacteria, chlorophyll auto-fluorescence for algae).
• Observation: Use light microscopy, fluorescence microscopy, or electron microscopy depending on the resolution and detail required.

DNA and RNA Analysis:

DNA Extraction:

RNA Extraction:

PCR and Sequencing:

• PCR Amplification: Use specific primers to amplify target regions of DNA or cDNA (for RNA). Perform PCR reactions under optimized conditions.
• Sequencing: Send purified PCR products for sequencing (Sanger or next-generation sequencing) to obtain genetic information.
• Bioinformatics: Analyze sequencing data using appropriate bioinformatics tools and software to identify and characterize microorganisms.

Metagenomics and Metatranscriptomics:

• Library Preparation: Prepare DNA or RNA libraries for high-throughput sequencing.
• Sequencing: Use next-generation sequencing platforms (e.g., Illumina, PacBio) for metagenomic or metatranscriptomic analysis.
• Data Analysis: Use bioinformatics pipelines for sequence assembly, annotation, and analysis to understand the microbial community structure and function.

Summary

The process of collecting, preserving, and analyzing microorganisms involves meticulous planning and execution to ensure sample integrity and reliable results. By following these steps, researchers can effectively study microorganisms from environmental samples and gain insights into their roles and functions in various ecosystems.

Dear Qasim,

Thank you so much for your complete and comprehensive answer.

Would you please introduce me to some papers about every level?

With best wishes and Good luck.

Jafar

Dear Qasim,

Hi,

Thanks for your kind efforts.

I have some questions about changing your advice to a more practical method for me or everyone:

1-For sterilizing bottles or containers could I use industrial alcohol?

2-At the sample collection level, the water sampling part, what mesh prefer you, to use a plankton net to concentrate the organisms?

3-At the Packaging and Transporting level, the Transport Conditions part: you suggest, maintaining them at 4°C during transport. Is it enough ice particles and an unolite box (Polystyrene container) for this level? I am confused about this degree with the Preservation Methods level; For DNA, freeze samples at -20°C; for RNA, freeze at -80°C. Is it 4 degrees to transport level Lab with ice? (I notice For Bacteria and Algae: DNA: I use commercial DNA preservation solutions stabilizer nucleic acids at room temperature.)

4-Immediate Processing: if I have preservation solutions available, can I do processing in-room temperature?

Best wishes,

Jafar

To collect microbial samples from environments to the laboratory:

4o mini

tanks a lot

Would you Please add here your private reply to the usage of other researchers?

• Dear Qasim, Hi, Thanks for your kind efforts. I have some questions about changing your advice to a more practical method for me or everyone: 1-For sterilizing bottles or containers could I use industrial alcohol? 2-At the sample collection level, the water sampling part, what mesh prefer you, to use a plankton net to concentrate the organisms? 3-At the Packaging and Transporting level, the Transport Conditions part: you suggest, maintaining them at 4°C during transport. Is it enough ice particles and an unolite box (Polystyrene container) for this level? I am confused about this degree with the Preservation Methods level; For DNA, freeze samples at -20°C; for RNA, freeze at -80°C. Is it 4 degrees to transport level Lab with ice? (I notice For Bacteria and Algae: DNA: I use commercial DNA preservation solutions stabilizer nucleic acids at room temperature.) 4-Immediate Processing: if I have preservation solutions available, can I do processing in-room temperature? Best wishes, Jafar
• 📷Qasim Ali to you, Asalam o aliakum While industrial alcohol can be used as a preliminary step for surface disinfection, it is not sufficient on its own for sterilizing containers for DNA or RNA isolation. Autoclaving, dry heat sterilization, or the use of specific chemical sterilants are recommended for thorough decontamination. The preferred mesh size for a plankton net largely depends on the target organisms of your study. For phytoplankton, use a fine mesh size (20-50 micrometers). For zooplankton, use a coarser mesh size (100-200 micrometers). For mixed plankton, an intermediate mesh size (50-100 micrometers) may be appropriate. Always consider your specific research goals and the characteristics of the sampling environment when selecting the mesh size. Maintaining samples at 4°C with ice packs and a polystyrene container is generally sufficient for short-term transport from the field to the lab. For DNA and RNA preservation, once samples reach the lab, they should be processed or stored at -20°C for DNA and -80°C for RNA. Using commercial preservation solutions can offer additional stability, particularly for sensitive samples like RNA. Always aim to minimize the time samples spend at non-optimal temperatures to ensure the highest quality for subsequent analyses. Using preservation solutions allows you to perform immediate processing of samples at room temperature with confidence in the stability of nucleic acids. These solutions provide an effective means to preserve the integrity of DNA and RNA until you are ready for extraction and further processing. Always follow the manufacturer’s guidelines for the best results and ensure compatibility with your extraction protocols. Good Luck

salam, Thank you very much, Dear Qasim I added your detailed reply in the question part, to other researcher's usage. I think to use a finer mesh size, had better because of the diatoms. Thank you for your kind reply. best wishes Jafar