N2Jenomics Lab Pvt. Ltd. offers comprehensive Degradome Sequencing (PARE) services to identify microRNA (miRNA)-mediated RNA cleavage sites and validate miRNA target genes on a genome-wide scale. Our integrated workflow combines high-throughput sequencing with advanced bioinformatics to provide accurate insights into post-transcriptional gene regulation, making it particularly valuable for plant functional genomics, although the approach can also support other RNA cleavage studies.

What Is Degradome Sequencing?

Degradome Sequencing, also known as Parallel Analysis of RNA Ends (PARE), is a high-throughput sequencing technique used to identify the cleavage products generated by miRNAs and other small regulatory RNAs. By sequencing the 5′ ends of degraded messenger RNAs (mRNAs), this approach enables genome-wide mapping of RNA cleavage sites and direct validation of miRNA-target interactions.

Compared with traditional 5′ RACE, Degradome Sequencing offers a faster, more sensitive, and scalable approach for identifying RNA cleavage events across the entire transcriptome.

This technology is widely used to investigate:

• • miRNA-target interactions

• • Post-transcriptional gene regulation
• • Small RNA regulatory networks
• • RNA degradation pathways
• • Functional genomics in plants

Advantages of Degradome Sequencing

• Genome-Wide miRNA Target Identification

Directly identifies cleavage sites of known and novel miRNA target genes across the transcriptome.

• Validation of Regulatory Networks

Supports the construction and validation of miRNA-mediated gene regulatory networks.

• High-Resolution Cleavage Site Mapping

Precisely maps RNA cleavage sites at nucleotide resolution, providing strong evidence for miRNA-target interactions.

• High-Throughput Analysis

Simultaneously analyzes thousands of transcripts in a single experiment, enabling comprehensive transcriptome-wide studies.

• Supports Functional Genomics

Provides valuable insights into gene regulation, developmental processes, stress responses, and disease mechanisms, particularly in plant research.

• Integration with Small RNA Sequencing

Can be combined with Small RNA Sequencing to identify novel miRNAs and experimentally validate their target genes for a more complete understanding of RNA-mediated regulation.

Applications

Degradome Sequencing is widely used for:

• • Genome-wide miRNA target identification
• • Validation of predicted miRNA targets
• • Plant functional genomics
• • Post-transcriptional gene regulation studies
• • Small RNA regulatory network analysis
• • Stress response and developmental biology research
• • Crop improvement and agricultural genomics
• • Comparative transcriptomics
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Degradome Sequencing Workflow

Our Degradome Sequencing (PARE) workflow is optimized to accurately identify miRNA-mediated RNA cleavage sites while ensuring high-quality, reproducible results.

1. RNA Preparation

High-quality poly(A)+ RNA is isolated from the sample and assessed for integrity and purity.

2. Library Construction

A specialized RNA adapter containing an MmeI recognition site is ligated to the 5′ ends of cleaved RNA fragments. The RNA is then reverse-transcribed to generate first-strand cDNA, followed by second-strand synthesis.

3. Library Processing

The double-stranded cDNA is digested with MmeI, size-selected through purification, ligated to a 3′ sequencing adapter, and PCR-amplified to generate the final degradome sequencing library.

4. High-Throughput Sequencing

The prepared libraries are sequenced on advanced next-generation sequencing (NGS) platforms to generate high-quality degradome data.

5. Bioinformatics Analysis

Sequencing data are processed to identify RNA cleavage sites, validate miRNA-target interactions, map degradome signatures, and construct regulatory networks.

Service Specifications

	Sample Requirements Sample type: Total RNA without degradation or DNA contamination Total RNA≥ 20 μg, Minimum Quantity: 15 μg, Concentration≥ 100 ng/µL 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 Degradome library preparation Illumina HiSeq SE50 More than 80% of bases with a ≥Q30 quality score
	Bioinformatics Analysis We provide multiple customized bioinformatics analyses: Raw data quality control Reference-based mapping Identification of known and novel non-coding RNAs Distribution analysis of degradation fragments on the selected region of genome Identification of target mRNAs Statistical summary of mRNA degradation site GO/KEGG analysis 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 Degradome Sequencing for your writing (customization)

1. What is the principle of Degradome Sequencing?

Degradome Sequencing, also known as Parallel Analysis of RNA Ends (PARE), identifies RNA molecules that have been cleaved by microRNAs (miRNAs) or other small regulatory RNAs. In plants, miRNAs typically cleave their target mRNAs between the 10th and 11th nucleotides of the complementary binding region, producing characteristic RNA degradation products. By sequencing the 5′ ends of uncapped RNA fragments, Degradome Sequencing enables genome-wide identification of miRNA-mediated cleavage sites and direct validation of miRNA-target interactions.

2. What are the advantages of Degradome Sequencing for miRNA target identification?

Compared with conventional validation methods, Degradome Sequencing offers several important advantages:

Because it combines high throughput, high sensitivity, and genome-wide analysis, Degradome Sequencing is an efficient approach for discovering and validating miRNA target genes.

3. How does Degradome Sequencing differ from other RNA sequencing methods?

Unlike conventional RNA-Seq, which measures transcript abundance across the entire transcriptome, Degradome Sequencing specifically analyzes 

• RNA degradation products. It captures the 5′ ends of cleaved RNA molecules, allowing researchers to directly identify • miRNA- and siRNA-mediated cleavage events and validate regulatory interactions.

While standard RNA-Seq reveals which genes are expressed, Degradome Sequencing reveals which transcripts are being actively cleaved and regulated by small RNAs, making the two approaches highly complementary.

4. Which organisms are best suited for Degradome Sequencing?

Degradome Sequencing is most widely used in plant research, where miRNAs primarily regulate gene expression through direct cleavage of target mRNAs. It is extensively applied in studies of plant development, stress responses, crop improvement, and functional genomics. Although the technique can also support specific RNA cleavage studies in other organisms, its greatest utility is in plant biology.

5. Can Degradome Sequencing be combined with Small RNA Sequencing?

Yes. Combining Small RNA Sequencing with Degradome Sequencing provides a comprehensive view of small RNA-mediated gene regulation. Small RNA Sequencing identifies known and novel miRNAs, while Degradome Sequencing experimentally validates their target genes by mapping RNA cleavage sites, enabling the construction of high-confidence miRNA regulatory networks.

6. What bioinformatics analyses are included in Degradome Sequencing?

A typical Degradome Sequencing analysis includes:

• • Raw sequencing data quality control
• • Adapter trimming and read filtering
• • Read alignment to the reference genome or transcriptome
• • Identification of RNA cleavage sites
• • miRNA target prediction and validation
• • Cleavage site categorization
• • Regulatory network analysis
• • Functional annotation
• • GO and KEGG pathway enrichment analysis (optional)