Full-length Transcriptome Sequencing

Introduction

Full-length transcriptome sequencing, based on the extraordinarily long reads available with the Nanopore or/and Pacbio platform, provides full-length transcripts information sets of specific tissues or organs of the species with or/and without reference genomes. Compared with next generation sequencing, Nanopore or/and Pacbio technology provides full-length reads spanning entire and high-quality transcript isoforms from the polyA-tail to the 5’ end without the need for assembly. It is ideal for improving annotations in reference genomes, characterizing gene isoforms, alternative splicing, alternative polyadenylation, and gene fusion events, as well as complementing the short / incomplete transcripts for species without a reference genome.

Data analysis Pipeline

 
Nanopore-pipeline

 
Pacbio-pipeline

Advantages and Application Areas

Study on Genome Structure

Alternative splicing, alternative polyadenylation, gene fusion events, and CDS prediction, etc.

 Gene Functional analysis

Identify full-length transcripts accurately. Based on gene functional annotation analysis, researchers could expand the complexity of known or novel transcription factors.

Improving genome annotation

Based on the third-generation sequencing, using full-length transcript isoforms can improve the accuracy of complex genome structure prediction.

Transcript expression analysis

Pacbio: Quantification assisted by next generation sequencing data

Nanopore: Directly Quantification

Quantification of gene/transcript expression levels could help researchers identify  differentially expressed genes and transcripts comprehensively and effectively.

Differential Structure Analysis of Multiple Samples Transcripts

Based on differential structure analysis of multiple samples, researchers could learn about specific or common transcripts at different developmental stages or under different treatments, revealing physiological mechanism.

Integrative analysis with proteome

Construct transcriptome-Proteomic Database; Verify with alternative splicing events mutually 


Biomarker Successful Cases

Biomarker firstly acquired Pacbio RSII and Sequel platforms in 2016, and then achieved long-term strategic cooperation with Nanopore Technologies in August 2018, which is the first and only one company ownned official certification with nanopore RNA sequencing in mainland China (updated in April, 2019). Until now, we have accumulated abundant project experience on both experiments and biological information levels, which is much higher than the average level of the market in China. There are many successful projects, such as Populus, sweet potato, rabbit, insects, Dolly Varden char and so on. In the end, Biomarker is one of the most powerful technology-service companies in the Third Generation Sequencing in China.

Projects Examples

[1] Identification of genes associated with ricinoleic acid accumulation in Hiptage benghalensis via transcriptome analysis[J]. Biotechnology for Biofuels, 2019, IF=5.497.

[2]Analysis of transcripts and splice isoforms in Medicago sativa L. by single-molecule long-read sequencing[J]. Plant Molecular Biology, 2019, IF=3.543.

[3]Comprehensive identifcation of the full-length transcripts and alternative splicing related to the secondary metabolism pathways in the tea plant (Camellia sinensis)[J]. Scientific Reports, 2019, IF=4.122.

[4]Full-Length Transcriptome Survey and Expression Analysis of Cassia obtusifolia to Discover Putative Genes Related to Aurantio-Obtusin Biosynthesis, Seed Formation and Development, and Stress Response[J].International Journal of Molecular Sciences, 2018, IF=3.687.

[5] The developmental dynamics of the Populus stem transcriptome[J]. Plant Biotechnology Journal, 2018, IF=7.443.

[6] SMRT sequencing of full-length transcriptome of flea beetleAgasicles hygrophila (Selman and Vogt):[J]. Scientific Reports, 2018, IF= 4.225 .

[7] Do the toll-like receptors and complement systems play equally important roles in freshwater adapted Dolly Varden char(Salvelinus malma)?[J]. Fish Shellfish Immunol, 2018, IF=3.185.

[8] Transcriptome profiling using Illumina- and SMRT-based RNA-seq of hot pepper for in-depth understanding of genes involved in CMV infection[J]. Gene, 2018, IF= 2.196 .

[9] A transcriptome atlas of rabbit revealed by PacBio single-molecule long-read sequencing[J]. Scientific Reports, 2017,IF= 4.225.

[10] Identification and analysis of glutathione S-transferase gene family in sweet potato reveal divergent GST-mediated networks in aboveground and underground tissues in response to abiotic stresses[J].BMC Plant Biology, 2017, IF=3.964.