What is 16S/18S/ITS Amplicon Sequencing?

16S/18S/ITS amplicon sequencing is a targeted next-generation sequencing (NGS) approach used to identify and profile microbial communities without the need for culturing. It enables accurate analysis of bacteria, archaea, fungi, and other eukaryotic microorganisms across a wide range of research applications.

At N2Jenomics Lab Pvt. Ltd., we deliver high-quality, cost-effective amplicon sequencing services to support studies in environmental science, agriculture, food safety, biotechnology, and biomedical research.

• 16S rRNA Sequencing

Targets the 16S rRNA gene to identify and classify bacterial and archaeal communities. Sequencing variable regions such as V3–V4 or V4–V5 enables reliable taxonomic profiling, making this method ideal for microbiome research, environmental monitoring, and microbial diversity studies.

• 18S rRNA Sequencing

Analyzes the 18S rRNA gene to profile eukaryotic microorganisms, including protists, algae, fungi, and other microeukaryotes. It is widely used for biodiversity assessment, ecological studies, and environmental microbiome analysis.

• ITS Sequencing

Targets the Internal Transcribed Spacer (ITS) region, the standard genetic marker for fungal identification. ITS sequencing provides high-resolution fungal profiling and is commonly used for mycobiome research, pathogen detection, biodiversity analysis, and food quality testing.

16S vs. 18S vs. ITS: Which Marker Is Right for Your Study?

Choosing the right amplicon marker is essential for obtaining accurate microbial profiling results. The comparison below can help you select the most suitable target based on your sample type and research objectives.

*Species-level classification varies by organism and reference database.

Quick Selection Guide

• • Choose 16S rRNA for bacterial and archaeal microbiome studies.

• • Choose 18S rRNA for profiling eukaryotic microorganisms such as protists and algae.
• • Choose ITS for high-resolution fungal identification and community analysis.

Why Choose Amplicon Sequencing?

Amplicon sequencing is a rapid and cost-effective approach for characterizing microbial communities without the need for culture-based methods.

• • Accurate Microbial Profiling – Reliable identification of diverse microbial populations.
• • High Sensitivity – Detects low-abundance organisms with deep sequencing coverage.
• • High-Resolution Analysis – ASV-based workflows improve taxonomic precision.
• • Cost-Effective – Targets specific genetic markers, reducing sequencing costs.
• • Fast Turnaround – Delivers high-quality data within a short timeframe.

Amplicon vs. Metagenomic Sequencing

Amplicon Sequencing Applications

16S rRNA Sequencing

Target: Bacteria and Archaea

Applications:

• • Gut microbiome research
• • Environmental microbiology
• • Soil and sediment analysis
• • Water quality monitoring
• • Food microbiology
• • Industrial bioprocess monitoring

18S rRNA Sequencing

Target: Eukaryotic microorganisms

Applications:

• • Aquatic ecosystem studies
• • Soil eukaryotic diversity
• • Marine microbiology
• • Environmental biodiversity assessment
• • Ecological research

ITS Sequencing

Target: Fungi

Applications:

• • Fungal community profiling
• • Mycobiome research
• • Plant pathogen identification
• • Food contamination testing
• • Agricultural and environmental studies

16S/18S/ITS Amplicon Sequencing Service Options

Choose the sequencing solution that best matches your research objectives.

• Standard Amplicon Sequencing

• - Targeted 16S, 18S, or ITS regions
• - Reliable genus- to species-level profiling
• - Cost-effective for routine microbiome studies

• Full-Length Amplicon Sequencing

• - Complete 16S, 18S, or ITS gene sequencing
• - Improved taxonomic resolution
• - Enhanced phylogenetic analysis and species identification

• Absolute Quantitative Amplicon Sequencing

• - Combines qPCR with NGS
• - Measures absolute microbial abundance
• - Ideal for quantitative microbiome and biomarker studies

Standard vs. Full-Length vs. Absolute Quantitative Amplicon Sequencing

Selecting the right amplicon sequencing approach depends on your research objectives. Whether you need cost-effective microbial profiling, enhanced taxonomic resolution, or precise microbial quantification, each method offers unique advantages.

Comparison of Amplicon Sequencing Options

Which Option Should You Choose?

• Standard Amplicon Sequencing

Choose this option if your goal is to compare microbial community composition, diversity, and taxonomic profiles across multiple samples in a cost-effective manner.

• Full-Length Amplicon Sequencing

Ideal for projects requiring greater taxonomic resolution and more confident species-level identification, particularly in complex microbial communities.

• Absolute Quantitative Amplicon Sequencing

Best suited for studies where true microbial abundance is important, such as evaluating treatment responses, microbial load changes, biomarker discovery, and longitudinal microbiome research.

End-to-End 16S/18S/ITS Sequencing Workflow

We provide a comprehensive, one-stop service covering the entire workflow—from sample preparation to data analysis—ensuring both data quality and research efficiency.

16S/18S/ITS Amplicon Sequencing Workflow

• Sequencing Platforms

• - Illumina MiSeq™
• - Illumina NovaSeq 6000™ (PE250)

• Effective Read Length

• • 200–250 bp after quality filtering and adapter trimming

• Supported Target Regions

• - 16S rRNA
• - 18S rRNA

ITS

• - ITS1
• - ITS2

• Custom Amplicons

• Custom primers and target regions available upon request.

• Accepted Sample Types

We support a wide variety of biological and environmental samples, including:

• • Fecal samples
• • Soil and sediment
• • Freshwater and marine water
• • Skin and oral swabs
• • Plant and animal tissues
• • Fermentation cultures
• • Purified DNA
• • Other environmental and biological specimens

Comprehensive Bioinformatics Analysis

Our standardized bioinformatics pipeline transforms raw sequencing data into accurate, publication-ready microbial insights.

• High-Resolution Sequence Processing

• - DADA2-based ASV generation
• - Single-nucleotide variant detection
• - High-confidence sequence filtering

• Taxonomic Classification

• - Microbial identification from phylum to species level*
• - Curated reference databases for improved annotation accuracy

*Species-level classification depends on the selected marker and reference database.

• Microbial Diversity Analysis

• - Alpha Diversity: Shannon, Chao1, Simpson, Observed Species
• - Beta Diversity: PCoA, NMDS, PCA, hierarchical clustering

• Differential Abundance Analysis

• - Taxonomic abundance profiles
• - Heatmaps and stacked bar charts
• - LEfSe biomarker discovery
• - Statistical comparison between sample groups

• Optional Absolute Quantification

• - Integration with qPCR-based workflows
• - Absolute microbial abundance estimation
• - Improved comparison across experimental groups

Research Questions Our Analysis Can Answer

Sample Submission Guidelines

Shipping Recommendations

• • Ship samples under refrigerated or frozen conditions.
• • Dry ice shipment is recommended for long-distance transport.
• • Protect samples from excessive heat and prolonged exposure during transit.
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Why Choose N2Jenomics Lab Pvt. Ltd.?

Our amplicon sequencing services combine advanced sequencing technologies with expert bioinformatics to deliver reliable microbiome insights for diverse research applications.

• High-Sensitivity Detection

Identify low-abundance microorganisms with deep sequencing coverage of 50,000+ reads per sample.

• Flexible Target Selection

Validated assays for 16S, 18S, ITS, and custom amplicon regions to support a wide range of study designs.

• Rapid Project Delivery

Efficient laboratory workflows and streamlined data analysis help shorten turnaround times.

• Publication-Ready Results

Receive comprehensive reports with high-quality visualizations, diversity analyses, biomarker identification, and statistical outputs suitable for scientific publications.

• End-to-End Scientific Support

Our experienced genomics and bioinformatics specialists provide guidance from experimental design through data interpretation, helping you maximize the value of your microbiome research.

1. How do I choose the right amplicon region for my project?

The optimal target region depends on your sample type and research objectives.

• • Gut microbiome: V3–V4 for balanced microbial coverage and taxonomic resolution.

• • Oral or skin microbiome: V1–V2 or V1–V3 for improved profiling of surface-associated bacteria.
• • Soil, water, and environmental samples: V3–V4 or V4–V5 for diverse microbial communities.
• • Fungal studies: ITS2 for general fungal profiling; ITS1 for plant- and air-associated fungi.
• • Eukaryotic microorganisms: 18S rRNA is recommended for protists and other microeukaryotes.

Our scientists can help you select the most suitable marker and primer set during project planning.

2. How many biological replicates are recommended?

Biological replicates improve the reliability of microbiome studies and statistical analyses.

• • Minimum: Three biological replicates per experimental group.
• • Recommended: Five to six replicates for highly variable or complex sample types.

3. What if my DNA sample does not meet the quality requirements?

Every sample undergoes quality assessment before sequencing. If DNA concentration or purity is insufficient, our team will recommend the best course of action, including optimized extraction or low-input workflows whenever possible.

4. Can you sequence challenging sample types?

Yes. We routinely process a wide range of sample types, including:

• • Soil, sediment, and sludge
• • Freshwater and marine water samples
• • Fecal samples and clinical swabs
• • Fermentation cultures
• • Low-biomass and complex environmental samples

Our workflows can be customized for difficult or contamination-prone specimens.

5. Is bioinformatics analysis included?

Yes. Our standard analysis package typically includes:

• • Raw data quality assessment
• • Taxonomic classification
• • Alpha and beta diversity analysis
• • Differential abundance analysis
• • Publication-ready reports and visualizations

• Advanced analyses, such as functional prediction, microbial interaction networks, pathway analysis, and customized statistical workflows, are available upon request.

6. Can I submit multiple samples in a single project?

Yes. We support high-throughput sequencing projects. Each sample is assigned a unique barcode to ensure accurate identification and prevent cross-sample contamination.

7. Do I need to provide primers?

No. We provide validated primer sets for commonly used targets such as 16S rRNA, 18S rRNA, and ITS regions. Custom primer design and optimization services are also available for specialized applications.

8. What is the difference between relative and absolute abundance?

Relative abundance measures the proportion of each microorganism within a sample, while absolute abundance estimates the actual microbial quantity or copy number. Absolute quantification provides a more accurate view of microbial population changes by accounting for differences in total microbial load.