5´UTR Sequencing Service: Mapping of Transcription Start Site Variants

Description

5´ UTR Sequencing Service for detecting 5´UTR Variants by constructing mRNA 5´-end enriched RNA-Seq NGS libraries

Reveal 5′ UTR Landscape and Transcription Start Site Usage genome-wide by 5´UTR-Sequencing

5′ End-Sequencing facilitates active transcript 5´ end detection, enrichment and mapping, and identifies known and novel transcription start sites (TSS) at single nucleotide resolution by next generation sequencing.

• Define 5′ UTR Landscape: Genome-wide detection of known and novel transcription start sites
• Single Nucleotide End Resolution: Transcription start sites are detected with single nucleotide resolution
• Identify Alternative Promoters: 5′ End-Sequencing detects alternative promoters for all endogenous genes
• High Throughput and Robust Workflow
• High Reproducibility with accurate data
• Unbiased with high specificity

The transcription start site (TSS) is the location of transcription initiation at the 5′ end of a gene sequence. Knowledge of the exact position of a TSS of an RNA molecule is crucial for the identification of regulatory regions immediately flanking it.
5´End-Seq facilitates precise UTR identification for RNA therapeutics targeting and can be used as a tool for identifying UTR biomarkers associated with disease. 5′ UTR Sequencing can quantify the relative usage of TSS across samples and indicates transcript isoform presence in annotated transcriptomes. Defining the 5′ UTR with 5´End-Seq can also help predict binding factor motifs more reliably.

5′ End-Seq Workflow
Total RNA is enriched for poly(A)+ mRNAs. Poly(A)+ mRNAs are heat fragmented and subsequently 5’ phosphorylated using PNK enzyme, which only uses uncapped molecules as substrate. The phosphorylated mRNA fragments are then digested by exonuclease, leaving behind only mRNA fragments with 5’-caps present. Those are used to prepare an RNA-seq library covering highly enriched sense-strand mRNA 5´ end fragments.

5′ End-Seq Service Description

You provide:

10 µg Total RNA; RIN ≥ 7; Non-Crosslinked

The service comprises:

• mRNA Enrichment
• RNA Fragmentation
• Exonuclease Digestion
• Adapter Ligation
• Library preparation
• Sequencing: 10-15M reads and comprehensice data analysis

Deliverable:

You receive the following comprehensive Data Analysis Package:

⋅ RAW SEQUENCING DATA | FASTQ:

Reads directly off sequencer and demultiplexed per sample for data back-up and publication

⋅GENOME ALIGNMENTS| BAM:

Reads filtered of repetitive elements, aligned, and PCR deduplicated for additional downstream analysis

⋅COVERAGE TRACKS| BIGWIG:

Normalized read end coverage on positive and negative strands for visualization in a genome viewer such as IGV

⋅SINGLE NUCLEOTIDE 5´ END CALLS| BED:

Genomic single nucleotide positions scored as putative 5’ transcript ends for 5’UTR exploration

-5′ End-Seq SUMMARY REPORT | HTML

Report with interactive tables and plots for publication ready graphics. Includes QC metrics and 5’ end enrichment

We are offering scaled pricing depending on the number of RNA samples provided –
please contact us for a quote!