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Full-length mRNA sequencing -PacBio
While NGS-based mRNA sequencing is a versatile tool for quantifying gene expression, its reliance on short reads restricts its use in complex transcriptomic analyses. On the other hand, PacBio sequencing (Iso-Seq) employs long-read technology, enabling the sequencing of full-length mRNA transcripts. This approach facilitates a comprehensive exploration of alternative splicing, gene fusions, and poly-adenylation. However, there are other choices for gene expression quantification due to the high amount of data required. PacBio sequencing technology relies on single-molecule, real-time (SMRT) sequencing, providing a distinct advantage in capturing full-length mRNA transcripts. This innovative approach involves using zero-mode waveguides (ZMWs) and microfabricated wells that enable the real-time observation of DNA polymerase activity during sequencing. Within these ZMWs, PacBio’s DNA polymerase synthesizes a complementary strand of DNA, generating long reads that span the entirety of mRNA transcripts. PacBio operation in Circular Consensus sequencing (CCS) mode enhances accuracy by repeatedly sequencing the same molecule. The generated HiFi reads have an accuracy comparable to NGS, further contributing to a comprehensive and reliable analysis of complex transcriptomic features.
Platform: PacBio Sequel II; PacBio Revio
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Eukaryotic mRNA Sequencing-NGS
mRNA sequencing is a technique that uses next-generation sequencing to reveal the presence and quantity of RNA molecules in a biological sample, providing a snapshot of gene expression in the sample.
With its wide-ranging applications, this cutting-edge tool unveils intricate gene expression profiles, gene structures, and molecular mechanisms associated with diverse biological processes.
This technique is widely adopted in research, clinical diagnostics, and drug development, since it offers insights into the intricacies of cellular dynamics and genetic regulation, providing a sturdy base to spark curiosity about its potential on different fields.
Platforms available: Illumina NovaSeq X; DNBSEQ-T7
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Non-Reference based mRNA Sequencing-NGS
mRNA sequencing empowers the comprehensive profiling of all mRNA transcripts within cells under specific conditions. This cutting-edge technology serves as a potent tool, unveiling intricate gene expression profiles, gene structures, and molecular mechanisms associated with diverse biological processes. Widely adopted in fundamental research, clinical diagnostics, and drug development, mRNA sequencing offers insights into the intricacies of cellular dynamics and genetic regulation.
Platform: Illumina NovaSeq X; DNBSEQ-T7
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Long Non-coding Sequencing-Illumina
Long non-coding RNAs (lncRNAs) are longer than 200 nucleotides that possess minimal coding potential and are pivotal elements within non-coding RNA. Found in the nucleus and cytoplasm, these RNAs play crucial roles in epigenetic, transcriptional, and post-transcriptional regulation, underscoring their significance in shaping cellular and molecular processes. LncRNA sequencing is a powerful tool in Cell differentiation, Ontogenesis, and Human diseases.
Platform: Illumina NovaSeq X
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Small RNA Sequencing-Illumina
Small RNA is a group of RNA molecules that include microRNAs (miRNAs), small interfering RNAs (siRNAs), and piwi-interacting RNAs (piRNAs). Among these, miRNAs, around 18-25 nucleotides long, are particularly noteworthy for their pivotal regulatory roles in various cellular processes. With tissue-specific and stage-specific expression patterns, miRNAs exhibit high conservation across different species.
Platform: Illumina NovaSeq
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CircRNA Sequencing-Illumina
Circular RNA sequencing (circRNA-seq) profiles and analyzes circular RNAs, a class of RNA molecules that form closed loops due to non-canonical splicing events, providing this RNA with increased stability. While some circRNAs have been shown to act as microRNA sponges, sequestering microRNAs and preventing them from regulating their target mRNAs, other circRNAs may interact with proteins, modulate gene expression, or have roles in cellular processes. circRNA expression analysis provides insights into the regulatory roles of these molecules and their significance in various cellular processes, developmental stages, and disease conditions, contributing to a deeper understanding of the complexity of RNA regulation in the context of gene expression.
Platform: Illumina Novaseq X
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Whole Transcriptome Sequencing – Illumina
Wole transcriptome sequencing offers a comprehensive approach to profiling diverse RNA molecules, encompassing coding (mRNA) and non-coding RNAs (lncRNA, circRNA, and miRNA). This technique captures the entire transcriptome of specific cells at a given moment, allowing for a holistic understanding of cellular processes.
Platforms availables: Illumina Novaseq X
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Chromatin Immunoprecipitation Sequencing (ChIP-seq)
Chromatin Immunoprecipitation (CHIP) is a technique that leverages antibodies to selectively enrich DNA-binding proteins and their corresponding genomics targets. Its integration with NGS enables the genome-wide profiling of DNA targets associated with histone modification, transcription factors, and other DNA-binding proteins. This dynamic approach enables comparisons of binding sites across diverse cell types, tissues, or conditions. ChIP-Seq’s applications span from studying transcriptional regulation and developmental pathways to elucidating disease mechanisms, making it an indispensable tool for understanding genomic regulation landscapes and advancing therapeutic insights.
Platform: Illumina NovaSeq
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Metagenomic Sequencing -NGS
A metagenome is a collection of the total genetic material of a mixed community of organisms, such as environmental and human metagenomes. It contains genomes of both cultivatable and uncultivatable microorganisms. Shotgun metagenomic sequencing with NGS enables the study of these intricate genomic landscapes embedded in environmental samples by providing more than taxonomic profiling, giving also granular insights into species diversity, abundance dynamics, and complex population structures. Beyond taxonomic studies, shotgun metagenomics also offers a functional genomics perspective, enabling the exploration of encoded genes and their putative roles in ecological processes. Finally, the establishment of correlation networks between genetic elements and environmental factors contributes to a holistic understanding of the intricate interplay between microbial communities and their ecological background. In conclusion, metagenomic sequencing stands as a pivotal instrument for unravelling the genomic intricacies of diverse microbial communities, illuminating the multifaceted relationships between genetics and ecology within these complex ecosystems.
Platforms: Illumina NovaSeq and DNBSEQ-T7
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Metagenomic Sequencing-TGS
A metagenome is a collection of the genetic material of a mixed community of organisms, such as environmental and human metagenomes. It contains genomes of both cultivatable and uncultivatable microorganisms. Metagenomic sequencing enables the study of these intricate genomic landscapes embedded in ecological samples by providing more than taxonomic profiling. It also offering a functional genomics perspective by exploring the encoded genes and their putative roles in environmental processes. While traditional shotgun approaches with Illumina sequencing have been widely used in metagenomic studies, the advent of Nanopore and PacBio long-read sequencing have changed the field. Nanopore and PacBio technology enhance downstream bioinformatic analyses, notably metagenome assembly, ensuring more continuous assemblies. Reports indicate that Nanopore-based and PacBio-based metagenomics have successfully generated complete and closed bacterial genomes from complex microbiomes (Moss, E. L., et al., Nature Biotech, 2020). Integrating Nanopore reads with Illumina reads provides a strategic approach for error correction, mitigating Nanopore’s inherent low accuracy. This synergistic combination leverages the strengths of each sequencing platform, offering a robust solution to overcome potential limitations and advancing the precision and reliability of metagenomic analyses.
Platform: Nanopore PromethION 48, Illumia and PacBio Revio
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Whole genome bisulfite sequencing(WGBS)
This technique, based on the bisulfite treatment of DNA to induce the conversion of unmethylated cytosines to uracil (C to U), while leaving methylated cytosines unchanged, stands as the gold-standard methodology for in-depth exploration of DNA methylation, specifically the fifth position in cytosine (5-mC), a pivotal regulator of gene expression and cellular activity.
This technique offers single-base resolution, allowing researchers to comprehensively investigate the methylome and uncover abnormal methylation patterns within the samples. By employing WGBS, scientists can gain unparalleled insights into genome-wide methylation landscapes, providing a nuanced understanding of the epigenetic mechanisms that underlie diverse biological processes and diseases.
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Assay for Transposase-Accessible Chromatin with High Throughput Sequencing (ATAC-seq)
ATAC-seq is a high-throughput sequencing technique used for genome-wide chromatin accessibility analysis. It use provides deeper understanding of the complex mechanisms of global epigenetic control over gene expression. The method uses a hyperactive Tn5 transposase to simultaneously fragment and tag open chromatin regions by inserting sequencing adaptors. Subsequent PCR amplification results in the creation of a sequencing library, which allows for the comprehensive identification of open chromatin regions under specific space-time conditions. ATAC-seq provides a holistic view of accessible chromatin landscapes, unlike methods that solely focus on transcription factor binding sites or specific histone-modified regions. By sequencing these open chromatin regions, ATAC-seq reveals regions more likely to active regulatory sequences and potential transcription factor binding sites, offering valuable insights into the dynamic modulation of gene expression across the genome.
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16S/18S/ITS Amplicon Sequencing-PacBio
The 16S and 18S rRNA genes, along with the Internal Transcribed Spacer (ITS) region, serve as pivotal molecular fingerprinting markers due to their combination of highly conserved and hyper-variable regions, making them invaluable tools for characterizing prokaryotic and eukaryotic organisms. Amplification and sequencing of these regions offer an isolation-free approach for investigating the microbial composition and diversity across various ecosystems. While Illumina sequencing typically targets short hypervariable regions like V3-V4 of 16S and ITS1, it has been demonstrated that superior taxonomic annotation is achievable by sequencing the full length of 16S, 18S, and ITS. This comprehensive approach results in higher percentages of accurately classified sequences, achieving a level of resolution that extends to species identification. PacBio’s Single-Molecule Real-Time (SMRT) sequencing platform stands out by providing highly accurate long reads (HiFi) that cover the full-length amplicons, rivalling the precision of Illumina sequencing. This capability allows researchers to attain an unmatched advantage — a panoramic view of the genetic landscape. The extended coverage significantly elevates the resolution in species annotation, particularly within bacterial or fungal communities, enabling a deeper understanding of the intricacies of microbial populations.
