Next Generation Sequencing (NGS)

Harrick Plasma → Applications → Biology & Biomedical → Next Generation Sequencing (NGS)

Next Generation Sequencing (NGS), or deep sequencing, provides researchers and clinicians with extensive genetic data with a multitude of applications. By fragmenting genetic materials and performing millions to billions of reads simultaneously, next generation sequencing is faster, more reliable, and lower cost than traditional sequencing methods. However, NGS applications can be limited by sample size and quality. In this article, learn how plasma treatment is used in conjunction with next generation sequencing to optimize results.

Sample Preparation

Effective sample preparation is integral to obtaining high-quality, accurate next generation sequencing results. While the type of genetic material used in next generation sequencing varies widely, often the initial goal is to isolate a homologues sample pool. Acquiring a sample set with high purity is essential for avoiding bias. This task can be incredibly difficult for clinical samples, where target cell availability is low. New sample preparation methods are needed to generate high quality, and pure NGS libraries with low input.  

One promising avenue for cell isolation or DNA/RNA purification are microfluidic devices fabricated by plasma treatment. Microfluidics offer precise control over cells and biomolecules. Using physical or chemical controls, researchers can generate pure sample pools with high accuracy. Custom microfluidic devices can be rapidly produces at a low cost using PDMS bonding via plasma treatment.

Oncology: Circulating Tumor cell (CTC) Isolation

An area of intense focus for next generation sequencing is oncology, where individual genetic sequencing may impact patient health most dramatically. Researchers are developing new methods for personalized diagnosis, prognosis, and drug discovery. A key step in many methods is isolating target cells from patient samples, often circulating tumor cells (CTCs). However, isolating CTC’s with enough genetic material for useful sequencing practices is difficult, particularly in early stage cancer. Microfluidic devices offer promising results for isolating small samples that can be used with next generation sequencing requirements. In several of these cases, on chip assays can be used in conjunction with NGS to provide even more substantial results.

• Centerfuge Chip [Che et al]
• Lateral Magnetophoresis [Cho et al]; [Kang et al]
• Droplet microfluidics [Liu et al]; [Wong et al]
• Microfluidic Ratchet (size and deformability) [Park et al]

In the following articles, microfluidic devices are constructed by PDMS bonding with Harrick Plasma cleaners. These microfluidic devices are designed to isolate CTCs that may later be used for next generation sequencing.

DNA (or RNA) Purification

Following DNA or RNA extraction from a target cell population, further purification steps may be necessary prior to NGS library generation. In several next generation platforms, a specific DNA or RNA size range is needed for effective sequencing. Again, the control offered by microfluidic devices over biomolecules lends itself to optimal NGS results. 

In the following articles, oligomers are sorted by size using microfluidic devices for use in next generation sequencing.

Biomolecule Immobilization

Immobilizing DNA or RNA on substrate surfaces is an important step in the development of next generation sequencing platforms, particularly sequencing by synthesis. In NGS flow cells, oligomers are positioned to hybridize with incoming genetic material, for cluster generation and analysis. Plasma treatment chemically modifies substrates through the introduction of polar functional groups. This hydrophilicity surface can either directly bind biomolecules or can facilitate deposition of coatings, such as silanization.  

In the following articles, plasma surface modification is used to immobilize or manipulate biomolecules for use in next generation sequencing.