Target enrichment offers several advantages over more comprehensive genomic profiling for a variety of scientific and applications. During this webinar, we explore the advantages and limitations of target enrichment, and discuss how NEBNext Direct®, a novel solution for selective enrichment of genomic targets, addresses these challenges..
NEBuilder HiFi: the next generation of DNA assembly – In this webinar, we will explore some of the latest applications for NEBuilder HiFi DNA Assembly. View full Q & A summary
In this webinar, Laurence Etwiller and Jennifer Ong discuss their recent publications, and how library preparation affects sequencing accuracy and variant calling. View full Q & A summary
qPCR reagents enable discoveries across a multitude of disciplines. Although they are ubiquitous, they are not all created equal – many fall short in multiplexed assays, limiting their utility in this important area. In the design and development of our Luna® qPCR portfolio, we took a systematic approach, creating a novel data visualization method that drove our development efforts. We also engineered a new, thermostable, WarmStart®reverse transcriptase (RT) that facilitates robust performance of the Luna One-Step RT-qPCR products, even in multiplex applications.
High fidelity polymerases are everywhere—but why would you need a high fidelity ligase? And what do we even mean by “fidelity” when we’re talking about ligation? NEB Scientist and ligase expert Greg Lohman discusses mismatch ligation by DNA ligases and the molecular diagnostics applications that depend on the use of high-fidelity DNA ligases like NEB’s new HiFi Taq DNA Ligase to detect single base differences in DNA*.
* intended for use in molecular diagnostics applications that depend on high fidelity nick ligation. This is not a substitute for T4 DNA Ligase and is not suitable for cloning applications or adaptor ligation/NGS library prep.
As NGS library prep throughput increases, DNA shearing has become a more significant bottleneck in the workflow. Current methods are challenged with instrumentation and consumables cost, lack of scalability, DNA damage and bias. In this webinar, we provide an overview of the current fragmentation strategies, and introduce a new enzymatic DNA fragmentation reagent that is fully integrated into the library prep workflow. This innovative method utilizes a simple protocol for a wide range of input amounts and GC content and yields reliable results.
During this webinar, we will discuss various options for analyzing data using pipelines specifically designed for the NEBNext Direct target enrichment approach, including open-source tools for the utilization of unique molecule identifiers (UMIs) for identification of PCR duplicate molecules and consensus error correction for improved variant detection. We will showcase the ability to analyze NEBNext Direct data using various solutions that allow the pipelines to be executed by anyone through user-friendly, 3rd party software interfaces, to production-grade pipelines to be implemented by a bioinformatician.
In this webinar you will learn how to increase editing efficiency by directly introducing Cas9 ribonucleoproteins (RNPs) to cells through electroporation or lipofection. Rapid sgRNA synthesis requiring only a single user-supplied ~55mer single-stranded DNA oligonucleotide is described. Methods for assessing genome editing efficiency will be discussed including T7-endonuclease I-based methods, sequencing-based methods, and in vitroCas9 digestion.
T4 DNA Ligase is a key enzyme in molecular biology, particularly useful for its high activity in the joining of multiple DNA fragments with complementary cohesive ends or blunt ends. We have recently developed a single-molecule sequencing method to characterize end-joining fidelity (discrimination against ligating mismatched overhangs) and bias (sequence preferences) for short cohesive ends in a high throughput manner. This method allows determination of the relative frequency of all ligation products with or without mismatches, the position-dependent frequency of each mismatch, and sequence-dependent biases in ligation efficiency. We have applied the fidelity and bias data to optimize design of Golden Gate-type assemblies, allowing selection of overhang sets with minimized mismatch potential and high efficiency. Use of this data has allowed highly efficient and accurate assembly of up to 24 fragments in one pot, with the flexibility to choose junctions within native coding sequences
NEBNext Direct Custom Ready Panels employ a novel hybridization-based enrichment approach to selectively enrich nucleic acid targets in a single-day workflow. We have designed and optimized probes specific to the full exonic content of human genes associated with cancer, neurological disorders, autism, cardiovascular disease and other conditions. These are designed, balanced, and pooled on a per gene basis, and can be combined into NEBNext Direct Custom Ready Panels, allowing rapid turnaround of specific custom gene subsets. Here, we present the ability to rapidly deploy custom gene panels across a variety of panel sizes and content, while maintaining high specificity, uniformity of coverage across target content, and sensitivity to detect nucleic acid variants from clinically relevant samples.
RNA studies have been expanding rapidly, on topics ranging from traditional gene-coding mRNA and structural rRNA to regulatory RNAs such as microRNA and long non-coding RNA. Various technologies have been developed to examine RNA abundance, however, RT-qPCR assays remain the most convenient and reliable approach for profiling RNAs of interest. Despite the prevalence of RT-qPCR, non-linear quantitation and sub-optimal robustness continue to be major issues. To enable fast and linear RNA detection, we developed a unique SuperMix for cDNA synthesis utilizing a novel thermostable reverse transcriptase. This optimized formulation delivers robust RNA detection with unmatched consistency and convenience.
Genotyping assays are essential in a variety of clinical and agricultural applications. While microarrays and endpoint PCR assays have been traditionally employed for these purposes, these methods lack contextual data of surrounding genomic regions, are limited to single nucleotide variants, and produce data with high false-positive calls. Target enrichment combined with next-generation sequencing is a compelling alternative that provides more complete genomic information along with the ability to discover newly introduced variants. However, the high cost and limited sample throughput of existing target enrichment strategies have made this approach largely inaccessible for genotyping workflows. During this webinar, we will discuss the NEBNext Direct Genotyping Solution, which enables highly multiplexed sample processing to provide a tractable and cost-effective targeted NGS method for human, plant, and animal genotyping applications
Exonucleases are enzymes that degrade different types of DNAs in specific ways, while endonucleases cleave at specific DNA structures or modifications. Both exo- and endonucleases are useful as molecular biology tools. In this webinar, we will review the activities of exonucleases and endonucleases in more detail, provide insight on how to choose the right exo- or endonuclease for various molecular biology applications, and explain how to use these reagents when developing new molecular biology workflows.