Small RNA Sequencing (sRNA-seq)

Small RNAs are short (<200 nt in length) and usually non-coding RNA molecules. There are several biologically relevant and functionally diverse classes of sRNA of specific sizes and produced by different, genetically separable pathways. Small RNAs include miRNA, siRNA, piRNA, snoRNA, snRNA, tsRNA, and srRNA. One of the most representative and significant functions of small RNA is RNA silencing (RNAi), in this case, endogenously expressed microRNA (miRNA) or exogenously derived small interfering RNA (siRNA) induces degradation of the complementary mRNA.

Overview

The innovation of large-scale, next-generation sequencing has exponentially increased knowledge of RNA biology, with regard to the diversity, abundance, and function of various RNA molecules. Small RNA-seq is a powerful tool for analyzing small RNAs such as miRNAs, siRNAs, and piRNAs in a single sequencing run, allowing the evaluation and discovery of novel small RNA molecules and the prediction of their functions. These RNA-seq methods have provided an even more complete characterization of small RNA and promised further applications.

Our technologies at single-base resolution allow for small RNA detection from very small amounts of cellular materials, which can help you detect pre-known small RNA, discover new small RNA, and examine all small RNA for differential expression in any sample. We generate small RNA sequencing libraries directly from total RNA and Capture the complete range of small RNAs, to understand the roles they play. this will provide you with a comprehensive and efficient approach to understand post-transcriptional regulation and to discover novel biomarkers.

Project Workflow

Bioinformatics Analysis Pipeline

In-depth data analysis:

• Length distribution and quantitative analysis of small RNA
• Differential expression analysis of small RNA
• Prediction of small RNA
• Discovery of novel biomarkers
• Coding-non-coding gene co-expression network (CNC network)
• GO annotation analysis
• KEGG pathway analysis
• Interactive network analysis

Sample Requirements

RNA sample (concentration ≥ 1 ng/uL, quantity ≥ 2 ug)

1.8 ≤ OD_260/280 ≤ 2.2, OD_260/230≥2.0, RIN ≥ 6.5, 28S:18S ≥ 1.0.

Please make sure that the RNA is not significantly degraded.

Sample storage: RNA can be dissolved in ethanol or RNA-free ultra-pure water and stored at -80°C. RNA should avoid repeated freezing and thawing.

Shipping Method: When shipping RNA samples, the RNA sample is stored in a 1.5 mL Eppendorf tube, sealed with sealing film. Shipments are generally recommended to contain 5-10 pounds of dry ice per 24 hours.

Deliverable: FastQ, BAM, coverage summary, QC report, custom bioinformatics analysis.

Demo Results

FAQ

• • Why are there specific requirements for sample extraction in small RNA sequencing?

    • When extracting total RNA, it is recommended not to use column-based reagents to avoid losing small RNA fragments. If providing small RNA samples directly, you can use specialized small RNA extraction kits for extraction.

• • What types of samples can be sequenced in your small RNA service?

    • We can perform small RNA library sequencing on a wide range of samples, including total RNA from animals, plants, microorganisms, small RNA fragments under 200 nucleotides, gel-purified small RNA samples, cell cultures, serum/plasma, tissue fluid, FFPE samples, RIP samples, exosomes, and more, as long as they meet our Sample Submission Requirements.

• • Why do experiment results sometimes contain degraded rRNA sequences?

    • Total RNA often experiences slight degradation, and natural degradation processes occur within living organisms. As a result, experiment data may contain minor mRNA degradation fragments. However, this proportion is typically low and depends on the quality of the total RNA samples.

• • What additional information do customers need to provide besides the samples for small RNA sequencing?

    • Customers should provide the genome of the relevant species and related information about exons, introns, and repeat elements. If the genome of the specific species is not available, information from a closely related species should be provided.

References:

1. McCormick KP, Willmann MR, Meyers BC. Experimental design, preprocessing, normalization and differential expression analysis of small RNA sequencing experiments. Silence. 2011, 2(1):1-9.
2. Sharma CM, Vogel J. Experimental approaches for the discovery and characterization of regulatory small RNA. Current opinion in microbiology. 2009, 12(5):536-46.
3. Burroughs AM, Ando Y, Aravind L. New perspectives on the diversification of the RNA interference system: insights from comparative genomics and small RNA sequencing. Wiley Interdisciplinary Reviews: RNA. 2014, 5(2):141-81.

miRNA (microRNA) Sequencing

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Bacterial Small RNA-Seq

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piRNA Sequencing

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siRNA Sequencing

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Pandora-Seq

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