Genomic workflows can vary dramatically especially within Next-Generation Sequencing (NGS) space which heavily depends on the type, scale and purpose of sequencing, so it is important to tailor workflows to your exact laboratory and project needs. In this article, Momin Shah, Sr. Genomic Application Specialist at Beckman Coulter Life Sciences, discusses what to consider when creating a custom genomic workflow, and how we can help.

Optimizing DNA and RNA extraction protocols for NGS

NGS technologies have revolutionized genomic and clinical research in recent years. These powerful tools are opening up new avenues of investigation into genetic diversity, metagenomics, and epigenetics.

From microbiome analysis to evolutionary biology, the broad diversity of applications makes customization of DNA or RNA extraction protocols essential to successfully fulfill the requirements of each individual project. For example, DNA extraction protocols for research applications are likely to have specific targets in terms of timeframes and the overall number of samples for processing, whereas isolation and purification protocols for research applications may need a degree of flexibility to adapt to changes in ongoing project work.

The workflow steps can also vary depending on the sample types, for example, RNA isolation protocols intended for use with tissue samples often involve an optimized homogenization and lysis which may require additional hardware to achieve the required shaking and incubation.

Whatever the project’s needs, designing a workflow for the efficient isolation and purification of DNA, RNA or both simultaneously begins by understanding the sample type and its state of preservation e.g., FFPE, storage buffer, temperature. It is also critical to understand the batch size, throughput, downstream application and the ultimate goals of the study.

Partnering with an experienced NGS instrument and reagent supplier at the very beginning of a new project can help to ensure that your nucleic acid extraction protocols are optimized from the outset to save time and costs down the line.

How can you customize an NGS workflow?

As stated above, understanding the requirements of a project is crucial to optimizing any workflow. Similarly for NGS, allowing the selection of the most appropriate instrumentation, reagents and consumables is critical. You should factor in the following key parameters when designing a workflow:

The scale of the sequencing operation

Sample numbers, the number of genes or panels (if using amplicons or target capture workflows) to be sequenced per sample, turnaround time and the project timeline itself are key factors to determine the scale of an operation.

It may seem obvious, but a large-scale project aiming to create a genomic DNA library for multiple whole genomes will require a high capacity workflow, whereas a sole researcher that is only looking at a specific gene—and not wanting the burden or cost of creating a genomic library—will have much lower throughput demands. The entire workflow—hardware, consumables, and reagents— should be carefully selected to match these demands.

Hardware requirements of workflows

Most genomic protocols will require some type of hardware tool in order to manipulate DNA or RNA when preparing samples. For example, magnetic bead-based techniques are widely used in DNA/RNA isolations, PCR cleanups and size selections, requiring a set-up with magnetic array stand or plate compatible with the labware format of the samples.

Additionally, devices such as thermocyclers, Peltier devices, and plate shakers are common too. There are also a variety of DNA purification protocols—from solution-based chemistries to ion exchange columns—each with their own labware and hardware requirements. Many of the reagents used in extractions or NGS library prep also need to be kept warm or cool, so it is important to include heating/cooling devices when designing a system for the workflow.

The downstream work after extraction and NGS will also have its own hardware needs such as qPCR for quantifications and or a sequencing device.

Consumables

Customizing workflows to align with extant laboratory practices necessitates an understanding of the current labware formats, the capacity of these sample containers, the volumes of reagents required (kit size), and the types of reagent vessels employed, as well as the desired output sample size. Moreover, ensuring an adequate inventory of these consumables is critical to minimize system downtime and achieve the requisite throughput for processing samples according to the scheduled timeline.

Library prep kits

It is imperative that a supplier demonstrates the capability to deliver adequate quantities of kits within a designated timeframe. For instance, a laboratory anticipating the processing of 100,000 samples annually necessitates more advanced logistical planning compared to a laboratory processing merely 6,000 samples per year. This is to ensure that procurement is executed well in advance to accommodate necessary shipping durations.

Automation

Implementing automation within genomic workflows significantly diminishes manual intervention and enhances operational efficiency. Collaboration with suppliers that offer highly adaptable and open-platform technologies enables the customization of instrument protocols to meet specific scientific needs. Furthermore, it is crucial for suppliers to ascertain the extent of automation required—whether it involves semi-automated systems or fully automated, walkaway solutions. Comprehensive automation facilitates the integration of additional devices and protocols and may evolve into a production-style workflow. In such configurations, plates are automatically received, processed, and forwarded to subsequent processes without human intervention.

How can you customize the software for automated NGS workflows?

Upon determining the appropriate instrumentation and automation level, it is critical to develop a user interface that enables operators to input specific run requirements efficiently. This interface facilitates the calculation of essential parameters by the instrument, including the minimum volume of each sample, the requisite volumes of reagents for initial gene amplification, and the methodology for pooling samples. To enhance customization, various scripting languages, such as JavaScript, are employed. These allow for the development of a tailored user interface comprising dialogue boxes, input tabs, buttons, text fields, check boxes, and tables, thereby ensuring precise alignment with the established workflow.

Beckman Coulter Life Sciences application support

In the dynamic realm of genomic research, reliable support is essential for the optimization and continual adaptation of extraction and NGS workflows in response to evolving requirements. The Application Support Team at Beckman Coulter Life Sciences excels in aiding scientists to identify and implement solutions that are ideally suited to their current and future needs. By offering expert guidance on instrumentation and hardware, as well as customizing programs, our team enables researchers to proficiently manage the complexities associated with NGS workflows, thereby ensuring confidence throughout their scientific endeavors.

Click here to see our automated solutions for NGS library prep or contact us to find out more.