NEB Scientific Posters
Your search returned 82 results.
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Increasing Sensitivity of Transcriptome Profiling in Prokaryotic and Eukaryotic Samples by Depleting Abundant RNAs
RNA-Seq is a widely used technology with a broad range of applications, including differential expression analysis and alternative splice forms identification, in normal and disease contexts as well as in developmental studies. The technology has been pushed to extremes of very low and degraded samples but still battles with the challenge of having a large dynamic range of transcript expression. Highly expressed transcripts with minimal biological interest can dominate readouts, masking detection of more informative low-abundance transcripts. Here, we present an improved method to enrich for RNAs of interest by eliminating abundant, typically unwanted, RNAs.
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Increasing Power to Detect Low-Frequency Variants Using Dual-Unique Molecular Indices
The use of Unique Molecular Identifiers (UMIs) has become increasingly popular – offering a multitude of advantages – particularly when paired with Unique Dual Indices (UDI). Two major factors affecting sequencing accuracy are 1) duplication, arising from PCR amplification of library molecules, and 2) errors introduced during library preparation and sequencing on the flow cell. UMIs, when incorporated into library preparation, can account for and mitigate the impacts of both of these factors. UMIs improved the accuracy of low-frequency variant detection by 25-200%.
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A Customizable Approach for Selective Removal of Abundant RNAs Enhances the Sensitivity of Transcript Detection Across Species
We developed a user-friendly web tool to enable custom depletion of any RNAs of interest. We used this web tool and depletion method to remove rRNA from total RNA of various species, including the mosquito Aedes aegypti as well as archaea Thermococcus kodakarensis and Pyrococcus furiosus. Additionally, we used this approach to target coding RNAs in human total RNA, and supplemented an existing anti-rRNA probe set for depletion of both rRNA and the selected coding RNAs. Using strand-specific RNA sequencing we measured depletion efficiency and transcript expression. We achieved high depletion efficiency (up to 99%) for all targeted RNAs across species, while maintaining transcript abundance of non-targeted RNA. This translated into an enrichment of RNAs of interest and an increased depth of sequencing coverage.
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Induro™, a Novel Reverse Transcriptase for Nanopore Direct RNA Sequencing with Significantly Improved RNA 5′ Coverage
Induro Reverse Transcriptase is an intron-encoded RT with superior performance in the cDNA synthesis reaction. Longer read lengths, higher percentage of full-length genes and transcripts, and improved 5′ coverage can be achieved with Induro RT without increasing error rates or decreasing mapping rates, enabling direct RNA sequencing on the Oxford Nanopore Technologies® platform.
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EM-seq™ enables accurate and robust methylation detection of cell free DNA and FFPE DNA sample types
NEBNext® Enzymatic Methyl-seq (EM-seq™) offers several improvements over traditional sodium bisulfite-based methylome analysis, owing – in large part – to the gentler, enzyme-based workflow. Less DNA damage enables longer reads with less sequencing depth. This poster summarizes the use of EM-seq with traditionally challenging sample types, cell-free DNA (cfDNA) and formalin-fixed, paraffin-embedded DNA (FFPE DNA).
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NEBNext® Ultra™ II FS DNA: A Robust Enzyme-based DNA Library Preparation Method Compatible with Plant Samples
Fragmentation is a bottleneck in the standard NGS workflow. The NEBNext® Ultra™ II FS DNA Library Prep Kit addresses this challenge with one-step enzymatic fragmentation, end-repair, and dA-tailing. Samples of plant tissue can be difficult to completely fragment without bias due to their molecular structure, but the Ultra II FS DNA kits enable robust library prep from Arabidopsis thaliana, Oryza sativa, and Zea mays samples.
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An E.coli Cell Lysate Based System for in vitro Protein Synthesis
The NEBExpress™ Cell-Free E. coli Protein Synthesis System is a high-performing, versatile and robust cell-free protein synthesis system developed by genetic engineering E. coli, optimizing a reaction buffer, and employing stringent manufacturing practices. This system was developed for coupled in vitro transcription and translation reactions for a variety of applications such as high throughput protein screening and engineering, as well as synthetic biology.
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An E.coli lysate-based system for in vitro Protein Synthesis
The NEBExpress™ Cell-Free E. coli Protein Synthesis System has been developed for coupled in vitro transcription and translation reactions resulting in high yields of proteins of various sizes (up to 230 kDa) and origins. A genetically engineered E.coli strain ensures stability of template DNA, RNA, and protein product. The Cell-free E.coli Protein Synthesis System is compatible with PURExpress Disulfide Bond Enhancer for better folding, and NEBExpress™GamS Nuclease Inhibitor for enhanced yield from linear templates. The reaction buffer formulation is compatible with SDS-PAGE (no acetone or TCA precipitation needed) and protein synthesis can be sustained for 10 hours at 37 °C or up to 24 hours at lower temperatures. Reproducible batches of lysate are produced, using highly stringent biomanufacturing processes and quality standards.
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Genome-wide profiling of nuclease protected domains reveals physical properties of chromatin (2019)
In metazoan cell nuclei, chromatin is functionally divided into transcriptionally active (euchromatin) or inactive (heterochromatin) regions. These heterochromatin regions constitute large chromatin domains that are in close contact with the nuclear lamina. Such lamina-associated domains (LADs) are thought to organize chromosomes inside the nucleus and are enriched for repressive histone modifications. Genome-wide profiling of heterochromatin, especially LADs, is often challenging and warrants a simpler and direct method. Here we developed a new method, Protect-seq, aimed at identifying regions of heterochromatin via resistance to nuclease degradation followed by next-generation sequencing. We performed Protect-seq on the human colon cancer cell line HCT-116 and observed overlap with previously curated LADs. We provide evidence that these protected regions are enriched for the repressive histone modification H3K9me3 and to a lesser extent H3K9me2 and H3K27me3. Moreover, the loss of H3K9me3 in human cells leads to an increase in chromatin accessibility. In sum, we demonstrate a novel technique to identify nuclease inaccessible regions of the genome and our data is consistent with the model that repressive chromatin domains are compacted and targeted to the nuclear lamina, likely via HP1 proteins, which act as scaffolds to maintain chromatin architecture.
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Genome filtering identifies species-specific DNA biomarkers for Mansonella perstans and Mansonella ozzardi, which enable differentiation of these closely related species and other co-endemic filarial parasites (2019)
Mansoneliasis is caused by infection with the parasites Mansonella perstans, M. ozzardi and M. streptocerca and is transmitted by insects such as biting midges and black flies. Immunosuppression caused by the parasitic infection may lead to worsening of other medical conditions. Mansoneliasis patients are often co-infected with multiple filarial parasites and anti-helminthic treatment is complicated. In this study, a bioinformatic filtering approach identified new diagnostic biomarkers, which were used to develop sensitive and species-specific LAMP assays that were validated on both patient and insect samples for point-of-care diagnostics.
