Next-generation sequencing (NGS) refers to high-throughput, massively parallel sequencing technologies for studying genetic variation associated with disease or other biological research. NGS improves upon chain termination sequencing, or Sanger sequencing, which revolutionized biological research, allowing researchers to determine genome sequences for the first time.

The primary difference between NGS and first-generation sequencing is throughput. NGS offers parallel sequencing of millions of small fragments of DNA per run. With conventional sequencing, or Sanger sequencing, only a few bases can be decoded per reaction using a time-consuming analysis. To give an example of the volume and throughput improvements, NGS can sequence an entire human genome in a single day, while the Sanger sequencing method that was used to decider the first human genome took more than a decade to decode. The massive improvement in throughput for NGS also led to a dramatic decrease in DNA sequencing costs.

All laboratory information management systems (LIMS) for NGS allow users to manage samples and associated data more easily and effectively. For any NGS workflow, a good LIMS can:

• Streamline compliance with HIPAA, CLIA, CAP, and other standards
• Visually track massive volumes of sample data
• Connect your NGS instruments
• Eliminate the need to search for slow and disconnected Excel data
• Automate metadata recording and reporting
• Organize boundless tasks
• Manage variant analysis for tertiary analysis

NGS is a complex scientific process that covers many steps, with QC checks along the way to ensure high-quality, consistent data. These steps include:

• Sample collection
• Sample preparation
• Library preparation
• Target enrichment
• Sequencing
• Data analysis and reporting

LIMS can connect all the instruments and software packages that are used across your NGS workflow, from liquid handlers to instrumentation to NGS data analysis software. For the sequencers specifically, the instrument generates a massive amount of NGS data, which gets stored and analyzed. The LIMS then finds the key results and presents them in a report.

There are a few common challenges that NGS laboratories face, many of which can be prevented by an Agilent product or solution.

• Challenge: Storing data in a reusable way. Solution: Agilent SLIMS is beneficial for searching and creating links between samples, procedures, and sample results, compared to common spreadsheet systems, such as Microsoft Excel. While Excel is easy to use and versatile, it is less desirable for retrieving information or structuring information.
• Challenge: Ensuring the sample quality is always up to certain level. Solution: Routine QC checks can ensure sample quality. Integrated and automated sample QC is ideal for efficiency and ease of use in an NGS workflow, as is the case with Agilent TapeStation systems fully integrated with Agilent SLIMS.
• Challenge: Library preparation is complex and error prone. Solution: Agilent SLIMS provides intuitive controls and user guidance to set up protocols more easily and quickly, while automation can reduce contamination, errors, and costs per sample. The Agilent Bravo is a flexible liquid handling platform that can automate your NGS sample preparation, freeing up time while generating consistent data.

To assure compliance, an NGS lab should meet the following standards, including: HIPAA, CLIA, ISO 13485, ISO/IEC 17025, and ISO 15189.

NGS is a strategy that is used to test many genes simultaneously to determine the order of nucleotides in entire genomes or targeted regions of DNA or RNA. NGS can be used to detect variants or mutations. For example, in cancer, NGS can be applied to a patient’s tumor that has been biopsied or removed.

Real-time polymerase chain reaction (PCR) is an easier and more forgiving technique with limited ability to multiplex. NGS allows analysts to simultaneously determine many gene sequences while revealing exact changes. However, NGS is a more technically demanding and costly process than PCR.

Quantitative polymerase chain reaction (qPCR) works best for low-throughput laboratories with low target numbers. NGS is more scalable and better suited for high-throughput sequencing, and it offers higher discovery power and higher sensitivity.

Decades after the invention of sequencing DNA, more than 10,000 NGS systems are installed globally. With next-generation sequencing now accepted as current-generation sequencing, labs are now facing the challenge to provide higher throughput and get more organized. Regulatory bodies also play a role, as they require laboratories to record more data and be fitted for long-term tenures. As such, there is a need for implementing a digital solution to manage the entire NGS workflow with consistency, while focusing on productivity.

With SLIMS, in addition to a broad NGS portfolio, Agilent provides a comprehensive software solution that manages and tracks all samples over the laboratory workflow. This includes quality control measures to be taken during sample collection, before sample use, NGS library QC, and during target enrichment. SLIMS also provides the sophisticated controls needed to ensure that data remain securely sorted and accessible for years to come.