Uncovering DNA Secrets: A Comparative Guide to Sanger, NGS, Third-Generation, and Hybrid Sequencing Techniques

What is the methodology for various DNA sequencing methods?

Methods for introducing DNA into microbes often result in disruption of genes, allowing study of the effects of their loss on phenotypes.

There are several methods for DNA sequencing, including:

1- Sanger sequencing: Also known as the chain-termination method, this method uses dideoxynucleotides to halt the elongation of DNA strands during replication, leading to fragments of different lengths that can be separated by gel electrophoresis. This technique was the first to be developed and is still widely used for sequencing short fragments of DNA.

2- Next-generation sequencing (NGS): NGS encompasses a range of techniques that allow high-throughput sequencing of DNA fragments. These methods include Illumina sequencing, which uses reversible terminators to sequence millions of fragments in parallel; Ion Torrent sequencing, which measures changes in pH as nucleotides are incorporated into a complementary strand; and Pacific Biosciences sequencing, which uses real-time monitoring of single DNA polymerases to sequence long DNA strands.

3- Third-generation sequencing: These techniques use single-molecule sequencing to sequence long DNA strands in real-time. Examples include Oxford Nanopore sequencing, which passes DNA through nanopores and measures changes in electrical current to identify nucleotides, and single-molecule real-time (SMRT) sequencing, which uses fluorescently-labeled nucleotides to measure DNA synthesis in real-time.

4- Hybrid sequencing: This approach combines two or more sequencing technologies to generate more accurate and complete sequence data. For example, long-read sequencing technologies like PacBio SMRT sequencing can be combined with short-read sequencing technologies like Illumina sequencing to correct errors and generate high-quality, phased genome assemblies.

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