Step 1: Sample Collection 

Testing begins with our custom sampling kits designed specifically for aquarium environments. Each kit contains everything you need to capture the microscopic world growing in your tank:

For Microbiome Analysis:

• A specialized filter housed in a protective plastic case that captures bacteria and archaea floating freely in your water column
• A sterile swab for sampling the biofilm communities that colonize submerged pipes, equipment surfaces, and other infrastructure

For tankDNA Analysis:

• A similar filter system captures environmental DNA (eDNA) from all sources – intact cells, microscopic organisms, tissue fragments, and dissolved genetic material shed by every living thing in your aquarium.

The sampling process is straightforward and non-disruptive to your aquatic ecosystem. Simply follow the included instructions to collect water through the filter and swab biofilm surfaces. These samples represent a genetic snapshot of your entire aquarium’s biological community.

Step 2: DNA Extraction

Once your samples arrive at our laboratory, we begin the crucial process of DNA extraction. Aquarium samples present unique challenges – they’re rich in contaminants that can interfere with genetic sequencing. Our extraction protocols have been specifically optimized to overcome these obstacles.

We use proven techniques that efficiently separate and purify DNA while removing inhibitory substances that could compromise downstream analysis. This specialized approach ensures we capture high-quality genetic material from even the most complex aquarium environments, from pristine reef tanks to heavily bioloaded systems.

Step 3: Targeted Gene Amplification

Microbiome Analysis: For bacterial and archaeal communities, we amplify a specific region of the 16S ribosomal RNA gene – the gold standard for microbial identification. We use the widely-adopted 515f and 806r primer pair, the same primers used by coral reef microbiome researchers worldwide. This standardization means your results can be directly compared with published scientific studies, giving you access to a wealth of comparative data about healthy and diseased coral reef systems.

tankDNA Analysis: For eukaryotic organisms (everything from microscopic algae to parasites and pests), we target the 18S ribosomal RNA gene using specific primers that amplify genetic material from plants, animals, fungi, and protists. This captures the full spectrum of eukaryotic life in your aquarium, including organisms too small to see or too rare to detect by traditional observation.

The amplification process uses polymerase chain reaction (PCR) to create millions of copies of these diagnostic gene regions, providing enough genetic material for comprehensive sequencing analysis.

Step 4: Library Preparation for Sequencing

Before sequencing, each amplified DNA sample must be prepared according to the requirements of our Illumina sequencing platform. This involves attaching specialized adapter sequences to both ends of every DNA fragment through a second PCR process using extended primers.

These adapters serve as molecular handles that the sequencing instrument recognizes and uses to process each fragment. We also add unique molecular barcodes to each sample, allowing us to sequence multiple samples simultaneously while keeping their data separate throughout the analysis pipeline.

Step 5: High-Throughput Sequencing

Your prepared samples are loaded onto an Illumina sequencing platform, where millions of DNA fragments are sequenced simultaneously. The instrument reads the genetic code of each fragment, generating thousands to tens of thousands of high-quality DNA sequences from your single aquarium sample.

This massively parallel approach allows us to detect even rare organisms that represent less than 0.1% of your aquarium’s total microbial community – species that would be impossible to find using traditional culturing methods.

Step 6: Bioinformatics Analysis and Species Identification

The raw sequencing data undergoes sophisticated computational analysis to transform millions of DNA sequences into meaningful biological information:

• Quality Control: We filter out low-quality sequences and remove potential contaminants to ensure reliable results.
• Species Identification: Each high-quality sequence is compared against comprehensive genetic databases:
  • Microbiome data is matched against the GreenGenes database of bacterial and archaeal 16S sequences
  • tankDNA data is compared against the Silva database, which contains ribosomal DNA sequences from eukaryotic organisms worldwide
• Taxonomic Classification: Sequences are assigned to specific taxonomic groups, from broad categories (like “Proteobacteria”) down to genus and sometimes species level, depending on the quality of the sequence and database coverage.
• Community Analysis: We quantify the relative abundance of different organisms and analyze the overall structure and diversity of your aquarium’s microbial ecosystem.

Ready to explore the hidden world in your aquarium? Order your sampling kit today and discover the science behind your tank’s success – or the reason for its problems.