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3' mRNA-seq is a quantitative, genome-wide transcriptomic technique based on the barcoding of the 3' untranslated region (UTR) of mRNA molecules. Unlike standard bulk RNA-seq, where short sequencing reads are generated along the entire length of mRNA transcripts, only the 3' end of polyadenylated RNAs are sequenced in 3' mRNA-seq.
RNA-Seq can also be used to determine exon/intron boundaries and verify or amend previously annotated 5' and 3' gene boundaries. Recent advances in RNA-Seq include single cell sequencing, bulk RNA sequencing, [6] 3' mRNA-sequencing, in situ sequencing of fixed tissue, and native RNA molecule sequencing with single-molecule real-time sequencing. [7]
Schematic overview of the MERCURIUS BRB-seq workflow where up to 384 samples can be barcoded and multiplexed per kit.. Bulk RNA barcoding and sequencing (BRB-seq) is an ultra-high-throughput bulk 3' mRNA-seq technology that uses early-stage sample barcoding and unique molecular identifiers (UMIs) to allow the pooling of up to 384 samples in one tube early in the sequencing library preparation ...
However, SAGE sampling is based on sequencing mRNA output, not on hybridization of mRNA output to probes, so transcription levels are measured more quantitatively than by microarray. In addition, the mRNA sequences do not need to be known a priori, so genes or gene variants which are not known can be discovered.
(See Central dogma of molecular biology). mRNA structure, approximately to scale for a human mRNA, where the median length of 3′UTR is 700 nucleotides. In molecular genetics, the three prime untranslated region (3′-UTR) is the section of messenger RNA (mRNA) that immediately follows the translation termination codon.
Sequencing technology platforms commonly used for RNA-Seq [77] [78] Platform Commercial release Typical read length Maximum throughput per run Single read accuracy RNA-Seq runs deposited in the NCBI SRA (Oct 2016) [79] 454 Life Sciences: 2005 700 bp 0.7 Gbp 99.9% 3548 Illumina: 2006 50–300 bp 900 Gbp 99.9% 362903 SOLiD: 2008 50 bp 320 Gbp 99. ...
It is important to distinguish the 5' and 3' UTRs from other non-protein-coding RNA. Within the coding sequence of pre-mRNA, there can be found sections of RNA that will not be included in the protein product. These sections of RNA are called introns. The RNA that results from RNA splicing is a sequence of exons. The reason why introns are not ...
The enzyme RNA polymerase II attaches to the template DNA strand and catalyzes the addition of ribonucleotides to the 3' end of the growing sequence of the mRNA transcript. [9] In order to initiate its function, RNA polymerase II needs to recognize a promoter sequence, located upstream (5') of the gene.