Next-Generation Sequencing (NGS) Workflow and Fundamentals of Sample Preparation

Next-generation sequencing (NGS) allows experts to extract information related to genetics from the collected sample much faster, more reliably, and at a low cost.

Getting the DNA ready for the sequencing procedure needs several preparations of library sequencing and a few steps that are completely dependent on the sample type and platform of NGS.

In this article, we cover the fundamentals of creating a sample for next-generation sequencing (NGS) and the current scenario of the NGS preparation market.

What is Next-Generation Sequencing?

Next-generation sequencing (NGS) is a high-power tool that helps healthcare experts in getting the required insight into the species at the epigenetic, genomic, or transcriptomic level.

Since the discovery of the method known as Ground-breaking Sanger in 1977, there have been constant advancements in sequencing technologies allowing substantial enhancements.

The potential applications of NGS in the field of healthcare research are evolutionary studies, metagenomics, and population genetics. The NGS applications in the field of medicine are prognostics, diagnostics, and studies related to genetic diseases.

In the field of biotech, the applications of NGS are agriculture, farming improvement, and food safety.

Based on the particular needs, experts can choose the sequencing platforms among several others. Based on the specific needs, it is possible to choose among multiple typical technologies, thus resulting in different capabilities of the sequencing and accordingly reading the lengths.

However, the workflow of NGS is divided into three phases: extraction of samples, library preparation, and analysis/sequencing.

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Common Type of Sequencing Procedures

Sample preparation methods are different from one another because the technology being used in every process has unique considerations.

The availability of new applications for the procedures of sequencing is constantly getting developed. Following are some common kinds of sequencing.

  • Whole Genome Sequencing

This sequencing style is used for determining the complete DNA sequence of the living organism at one time. Nearly, such a biological sample having the complete DNA copy can allow getting the required genetic materials for complete genome sequencing.

  • Whole Exome Sequencing

This sequencing style mainly focuses on scanning the genetic variants which can change the protein sequences and can cause diseases. It is necessary to select the exons before they are sequenced; this method is known as targeted strategies.

How does Next-Generation Sequencing Work?

  • Sample Extraction

The NGS procedure starts with extracting the acids like nucleic acids, which are used to sequence DNA, mRNA, chromatin, or RNA.

Depending on the goal of the experiment, the genetic materials can be easily extracted from several biological samples like cultured cells, tissues sections, blood, urine, and plant specimens or microorganisms.

The quantity and quality of these extracted samples are essential for successful NGS. Therefore, it is highly mandatory to choose an appropriate method of extraction and establish quality parameters before moving to the next steps.

  • Library Preparation

At this step of NGS sample preparation, different processing steps are mandatory for generating a library.

In simple words, it can also be described as the step for transforming the extracted sample in the most suitable format as per the chosen technology for sequencing.

The samples of DNA are fragmented in uniform pieces by enzyme digestion or physical shearing. The ends of the resulting DNA fragments are linked to the technology for adaptor sequences.

This process helps in mixing multiple samples and getting sequenced at the same time, thus reducing the overall cost of the procedures.

  • Sequencing

Most of the technologies used for sequencing are completely based on detecting the nucleotides included by DNA polymerase, hence defined as “sequencing by synthesis” (SBS) methods.

The most common method used for sequencing is reversible terminator, pyrosequencing, and semiconductor sequencing.

The first phase starts with scanning the nucleotides present in each sequencing read; this phase is called base calling. Then, these reads are aligned as per the genome. Finally, the last step is to identify DNA variants and get the results.

Current Insight of NGS Sample Preparation Market

NGS sample preparation market growth is supposed to grow at a high pace because of the increasing usage of NGS in medical laboratories.

Advanced methods of genetic diagnosis are used for prenatal diagnosis and genetic testing. As the number of patents related to genetic disorders is getting high, the consistency and efficiency of the screening testing have become more important. The faster result results to having minimum errors in performing effective screening of the genetic disorders.

Some challenges restricting the NGS sample preparing market growth are the huge cost of the NGS sample instruments, lack of skilled professionals, and also infrastructure.

The value of the NGS sample preparation market in 2020 was $1,468.9 million. As per BIS Research, the market is projected to grow and touch $3,279.3 million by 2026, at a CAGR of 14.24% during 2021–2026.


Next-generation sequencing has been there for over two decades. The impact of advanced NGS simple preparation can help in altering the healthcare sector. It can be highly helpful in scanning genetic disorders and taking the right steps.

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BIS Research

BIS Research

BIS Research offers exclusive market intelligence reports on disruptive technologies. Each year we publish over 200 titles across industry verticals.

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