Difference Between Shotgun Sequencing and Next Generation Sequencing

The key difference between shotgun sequencing and next generation sequencing is that shotgun sequencing is a sequencing method which randomly breaks up DNA sequences into many small fragments and reassembles the sequence by observing the overlapping regions while next Next Generation Sequencing (NGS) is an advanced method of genetic sequencing which depends on capillary electrophoresis.

Sequencing is the process that determines the precise order of nucleotides in a gene, a cluster of genes, chromosome, and a complete genome. It is very important in genomic studies, forensic studies, virology, biological systematic, medical diagnosis, biotechnology and in many other fields to analyze the structure and function of genes and identification of organisms. Furthermore, there are different types of sequencing methods available. Shotgun Sequencing and Next Generation Sequencing are two advanced methods among them.

CONTENTS

1. Overview and Key Difference
2. What is Shotgun Sequencing 
3. What is Next Generation Sequencing
4. Similarities Between Shotgun Sequencing and Next Generation Sequencing
5. Side by Side Comparison – Shotgun Sequencing vs Next Generation Sequencing in Tabular Form
6. Summary

What is Shotgun Sequencing?

Shotgun sequencing is a sequencing method which randomly breaks DNA sequences of the entire chromosome or entire genome into many small fragments and reassembles the sequences by computers through observing the overlapping sequences or regions. Generally, mammalian genomes are structurally complex and larger in size. Hence, they are difficult to sequence by cloning since it is time-consuming. Shotgun sequencing is a faster method. It is also cheaper to carry out.  Therefore, modern-day scientists rely on shotgun sequencing method to tackle complex genomes.

Shotgun sequencing procedure is comparatively simple. It starts with fragmenting the entire genome into varying sizes from 20-kilo bases to 300-kilo bases. Then each and every fragment should be sequenced using the chain termination method. After sequencing, it is necessary to assemble the fragments by looking at the overlapping regions using sophisticated computer software. Conventional mapping and cloning of sequences are not necessary for this method. Furthermore, the use of genetic map doesn’t take place in this method. However, since there is no use of existing genome maps, errors during the assemblage are more likely to occur. It is one of the major disadvantages of this method. Furthermore, shorter fragments provide less unique information for each read in this method. Moreover, shotgun sequencing fails to produce enough data to determine a consensus sequence at the required standard of accuracy.

Despite having the above-mentioned drawbacks, the shotgun sequencing method is currently the most efficient and cost-effective strategy for sequencing microbial genomes, including bacteria, viruses, and yeast. It is because their genomes lack repetitive regions which are difficult to sequence, and it is possible to assemble these genomes easily into chromosomes without errors.

What is Next Generation Sequencing?

Next Generation Sequencing (NGS) is a term used to refer to modern high throughput sequencing processes. It describes a number of different modern sequencing technologies which revolutionized genomic studies and Molecular Biology. These techniques include Illumina sequencing, Roche 454 sequencing, Ion Proton sequencing and SOLiD (Sequencing by Oligo Ligation Detection) sequencing. NGS systems are quicker and cheaper. Four main DNA sequencing methods are used in NGS systems: pyrosequencing, sequencing by synthesis, sequencing by ligation and ion semiconductor sequencing. A large number of DNA or RNA strands (millions of) can be sequenced in parallel by NGS. It allows sequencing of the entire genome of organisms within a short time period.

NGS has different advantages. It is a high speed, more accurate and cost-effective process which can be performed with a small sample size. Hence, it enables the analysis of the entire human genome in a single sequencing experiment. Furthermore, NGS can be used in metagenomic studies, in the detection of variations within an individual genome due to insertions and deletions, etc., and in the analysis of gene expressions. Moreover, NGS can analyze entire transcriptomes from large numbers of tissues simultaneously. Hence, NGS has revolutionized the analysis of transcriptomes.

What are the Similarities Between Shotgun Sequencing and Next Generation Sequencing?

• Shotgun sequencing and next generation sequencing are two genome sequencing methods.
• Both methods are rapid methods.
• Moreover, they are cost-effective methods.
• They are able to sequence many DNA fragments in parallel.

What is the Difference Between Shotgun Sequencing and Next Generation Sequencing?

Summary – Shotgun Sequencing vs Next Generation Sequencing

Shotgun sequencing and next generation sequencing are two sequencing methods used in genome sequencing. Both methods are fast and cost-effective methods. NGS works on the principle of sequencing millions of sequences simultaneously in a rapid way through a sequencing system. In contrast, shotgun sequencing requires breaking of genomes into small fragments and sequencing and reassembling using overlapping sequences. So, this summarizes the difference between shotgun sequencing and next generation sequencing.

Reference:

1. “Complex Genomes: Shotgun Sequencing” Nature News, Nature Publishing Group, Available here.
2. Behjati, Sam, and Patrick S Tarpey. “What Is next Generation Sequencing?” Archives of Disease in Childhood. Education and Practice Edition, BMJ Publishing Group, Dec. 2013, Available here.

Image Courtesy:

1. “Whole genome shotgun sequencing versus Hierarchical shotgun sequencing” By Commins, J., Toft, C., Fares, M. A. – “Computational Biology Methods and Their Application to the Comparative Genomics of Endocellular Symbiotic Bacteria of Insects.” Biol. Procedures Online (2009). Accessed via SpringerImages (CC BY-SA 2.5) via Commons Wikimedia
2. “HiSeq 2000” By RE73 – Own work (CC BY-SA 3.0) via Commons Wikimedia