Hello Biostars community,

First of all, my apologies for asking many questions step by step here. I don’t have direct guidance, so I am relying on this platform to learn and improve. Thank you for your patience.

I am currently working on metagenomic data from rhizospheric soil, and my focus is on both prokaryotes and eukaryotes. At the moment, I am running assemblies and trying to figure out the best parameters and tools to obtain a good assembly.

After assembly, my plan is to perform taxonomic classification using both:

Assembly-based approach - to annotate MAGs.

For prokaryotes, I am considering GTDB-Tk.

For eukaryotes, I am exploring tools like Tiara and MetaEuk (still searching for the best options for eukaryotic MAGs).

Read-based approach - here I am very confused. There are k-mer-based, marker gene-based, and protein-level methods, but I am not sure which would be the most suitable to capture both prokaryotes and eukaryotes.

My main doubts:

1. Which methods are generally recommended for read-based taxonomic classification when both prokaryotes and eukaryotes are present?

2. Which databases are considered reliable for covering both domains?

3. Are there tools/workflows that people have found particularly effective for handling whole metagenomes (prokaryotes + eukaryotes)?

If anyone here has experience with such datasets, I would be very grateful for your guidance or suggestions on how to structure this pipeline.

Thank you in advance for your time and help.