N2Jenomics Lab Pvt. Ltd. offers comprehensive Total RNA Sequencing (Total RNA-Seq) services for unbiased transcriptome profiling across bacteria, plants, animals, and human research samples. Our optimized workflows combine efficient rRNA depletion, high-throughput next-generation sequencing, and advanced bioinformatics to simultaneously profile coding and non-coding RNAs, providing a complete view of the transcriptome.

Total RNA-Seq is ideal for studies investigating gene regulation, non-coding RNAs, transcript discovery, and complex transcriptional landscapes.

What Is Total RNA Sequencing?

Total RNA Sequencing is an RNA-Seq approach that analyzes both protein-coding transcripts (mRNA) and non-coding RNA (ncRNA) after the removal of abundant ribosomal RNA (rRNA). Unlike poly(A)-enriched RNA-Seq, Total RNA-Seq captures both polyadenylated and non-polyadenylated RNA molecules, enabling more comprehensive transcriptome analysis.

This approach supports the identification of:

• • Protein-coding transcripts (mRNA)

• • Long non-coding RNAs (lncRNA)
• • Small nuclear RNAs (snRNA)
• • Small nucleolar RNAs (snoRNA)
• • Circular RNAs (circRNA)
• • Other regulatory non-coding RNAs

• Total RNA-Seq also enables the detection of novel transcripts, alternative splicing events, gene fusions, allele-specific expression, and transcript variants, making it a versatile solution for functional genomics and biomarker discovery.

Advantages of Total RNA Sequencing

• Comprehensive Transcriptome Coverage

Simultaneously profiles both coding and non-coding RNA species for a complete overview of gene expression.

• Broad RNA Species Detection

Captures polyadenylated and non-polyadenylated transcripts, including lncRNAs, snRNAs, snoRNAs, and other regulatory RNAs.

• High-Quality rRNA Depletion

Optimized ribosomal RNA depletion maximizes informative sequencing reads and improves transcriptome coverage.

• Suitable for Diverse Sample Types

Compatible with high-quality RNA as well as challenging samples, including partially degraded and FFPE-derived RNA.

• Novel Transcript Discovery

Identifies previously unannotated transcripts, alternative splice variants, gene fusions, and transcript isoforms.

• Allele-Specific Expression Analysis

Supports the detection of allele-specific gene expression and transcript-level variation.

• Reliable and Cost-Effective

Provides accurate, reproducible transcriptome profiling with flexible sequencing depth and rapid turnaround times.

Applications

Total RNA Sequencing is widely used for:

• • Whole-transcriptome profiling
• • Coding and non-coding RNA analysis
• • Long non-coding RNA (lncRNA) research
• • Gene expression analysis
• • Differential gene expression studies
• • Alternative splicing analysis
• • Novel transcript discovery
• • Gene fusion detection
• • Allele-specific expression analysis
• • Biomarker discovery
• • Functional genomics
• • Disease mechanism research

Total RNA Sequencing Workflow

Our comprehensive RNA-Seq services provide the RNA sequencing workflow from sample preparation through data analysis, enable rapid profiling and deep insight of the RNA.

Service Specifications

	Sample Requirements Total RNA ≥ 2 μg, Minimum Quantity: 500 ng, Concentration≥ 50 ng/µL Cells≥ 2×106 Tissue≥ 500 mg, Minimum Quantity: 100 mg A260:A280 value should be between 1.8 and 2.0 OD A260/A280 ratio ≥ 1.8, A260/230 ratio≥ 1.8, RIN ≥ 6 All total RNA samples should be DNA-free RNA should be stored in nuclease-free water or RNA Stable. Note: Sample amounts are listed for reference only. For detailed information, please contact us with your customized requests.
	Sequencing Strategy 250~300 bp insert cDNA library Illumina HiSeq 150 bp paired end Sequencing data size: 10 Gb >40 million reads
	Bioinformatics Analysis We provide multiple customized bioinformatics analyses: Data preprocessing Map to reference genome Small/Circular RNA identification Sequence analysis Differential expression analysis miRNA target gene analysis GO enrichment analysis KEGG enrichment analysis …and more Note: Recommended data outputs and analysis contents displayed are for reference only. For detailed information, please contact us with your customized requests.

Analysis Pipeline

Deliverables

• • The original sequencing data
• • Experimental results
• • Data analysis report
• • Details in Total RNA Sequencing for your writing (customization)

1. Should I choose Total RNA Sequencing or mRNA Sequencing?

The choice depends on your research objectives.

• • Total RNA Sequencing provides comprehensive transcriptome profiling by analyzing both coding (mRNA) and non-coding RNAs after ribosomal RNA (rRNA) depletion. It is ideal for studies involving lncRNAs, circRNAs, regulatory RNAs, transcript discovery, and whole-transcriptome analysis.

• • mRNA Sequencing enriches for polyadenylated (poly(A)) transcripts, making it the preferred option for studies focused on protein-coding gene expression, differential gene expression (DEG), and pathway analysis. It is also a more cost-effective approach when non-coding RNAs are not the primary interest.

Our scientific team can help determine the most suitable workflow based on your samples and research goals.

2. Which sequencing platforms are used for Total RNA Sequencing?

Total RNA Sequencing is primarily performed using Illumina short-read sequencing platforms, which provide highly accurate and cost-effective transcriptome profiling. For applications requiring complete transcript structures or isoform characterization, long-read sequencing technologies can be incorporated to complement short-read data.

3. What is the advantage of strand-specific Total RNA Sequencing?

Strand-specific library preparation preserves the original transcriptional orientation of RNA molecules, enabling more accurate transcriptome analysis. This approach improves:

• • Gene expression quantification
• • Genome and transcript annotation
• • Identification of antisense transcripts
• • Alternative splicing analysis
• • Novel transcript discovery
• • Characterization of overlapping genes

• Maintaining strand information is particularly valuable for studies involving complex genomes and non-coding RNAs.

4. How does Total RNA Sequencing support gene regulation and functional genomics research?

Total RNA Sequencing provides a comprehensive view of the transcriptome by simultaneously profiling coding and non-coding RNAs. This enables researchers to investigate:

• • Gene expression regulation
• • Alternative splicing events
• • Long non-coding RNA (lncRNA) function
• • Regulatory RNA networks
• • Gene fusion events
• • Novel transcript discovery
• • Cellular responses to developmental, environmental, or disease conditions

When integrated with genomic, epigenomic, or proteomic data, Total RNA-Seq offers deeper insights into molecular pathways, regulatory mechanisms, and biological processes.

5. Can Total RNA Sequencing detect non-coding RNAs?

Yes. Unlike mRNA Sequencing, Total RNA Sequencing captures both polyadenylated and non-polyadenylated RNA molecules, allowing comprehensive profiling of a wide range of non-coding RNAs, including lncRNAs, snRNAs, snoRNAs, circRNAs, and other regulatory RNA species. This makes it an excellent choice for studies focused on RNA biology and gene regulation.

6. Is Total RNA Sequencing suitable for degraded or FFPE samples?

Yes. Total RNA Sequencing with rRNA depletion is generally more suitable than poly(A)-based methods for partially degraded RNA, including many formalin-fixed paraffin-embedded (FFPE) samples. Because it does not rely solely on intact poly(A) tails, it can recover a broader range of transcripts from challenging samples while maintaining comprehensive transcriptome coverage.