mRNA Sequencing Service

What is mRNA Sequencing

mRNA is transcribed from protein-coding genes and serves as the intermediate RNA molecule that transfers genetic information from DNA to proteins. In eukaryotes, DNA is initially transcribed into pre-mRNA, which undergoes a series of processing steps to form mature mRNA. The mature mRNA is then transported to the cytoplasm and translated by ribosomes into proteins. The maturation of mRNA involves three main processing events: splicing, 5' capping, and 3' polyadenylation. As a result, mature mRNA molecules possess a poly(A) tail at their 3' end.

mRNA-seq is a high-throughput sequencing technology developed for detecting mRNA expression and processing. The study of mRNA sequencing in eukaryotes focuses on the collection of mRNAs transcribed from specific cells or tissues under certain functional states. It can be used to investigate differential mRNA expression, detect structural variations, and screen molecular markers associated with diseases or traits. It can also reveal the complexity of the transcriptome, determine gene and transcript structures, variable splicing, RNA editing, polyadenylated non-coding RNAs, and novel transcripts. Currently, mRNA-seq has been widely applied in fields such as basic research, molecular breeding, clinical diagnosis, and drug development.

mRNA-seq Technical Principle

For RNA containing poly(A) tails, such as mRNA and some lncRNAs, enrichment is performed using oligo dT, which captures the polyadenylated RNA, also known as PolyA-Seq. The enriched mRNA/lncRNA is then fragmented, reverse transcribed, ligated with common sequencing adapters, and PCR amplified to generate sequencing libraries. The resulting sequencing data is subjected to bioinformatics analysis to obtain information on gene expression, alternative splicing, RNA editing, and other features present in the RNA.

mRNA-seq Service

For mRNA-Seq, our company provides experimental services including but not limited to library construction and sequencing based on tissues and cell lines, as well as streamlined and customized bioinformatics analysis and interpretation services. The bioinformatics analysis services mainly include quality control and filtering of raw sequencing data, alignment of filtered data to a reference genome, mRNA quantification, differential gene analysis between different groups, sample clustering analysis, functional enrichment analysis of differentially expressed genes, protein interaction analysis of differentially expressed genes, and structural analysis of interaction networks. The core analysis results are as follows:

Differential gene analysis

Functional annotation and enrichment analysis

Protein interaction network and structural analysis

mRNA Sequencing Data Analysis

Advantages of mRNA Sequencing

Compared to gene expression microarrays, mRNA-Seq has several advantages in transcriptome analysis:

It has a broader dynamic range, increasing both sensitivity and accuracy in measuring fold changes in gene expression.

It can capture known features as well as novel features.

It can be widely applied to various species.

Our mRNA Sequencing Features

In addition to calculating gene expression levels, it can detect sequence and structural variations in transcripts.

Increasing sequencing depth allows for a wider dynamic range of detection, enabling the identification and quantification of both highly abundant and low abundant transcripts within a span of six orders of magnitude.

In addition to detecting the expression levels of known transcripts, it can also discover novel transcripts.

When applied to species without a reference genome, it can discover new genes through de novo assembly.

Service Specification

	Sample requirements and preparation Sample type: Total RNA without degradation or DNA contamination Starting amount of total RNA ≥ 5 µg Sample concentration ≥ 200 ng/µl Sample purity: OD260/280 = 1.8~2.2 All RNA samples are validated for purity and quantity
	Sequencing Libraries of miRNA (15-30nt), small RNA (30-200nt), or custom size range Illumina HiSeq SE50 ≥ 10 M reads More than 90% of bases with a ≥Q30 quality score