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Sanger Sequencing

Sanger Sequencing

DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...

RNA-seq

RNA-seq

RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases.
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...

Next-generation Sequencing

Next-generation Sequencing

The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
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Maxam-Gilbert Sequencing

Maxam-Gilbert Sequencing

In the same year as the discovery of the Sanger sequencing method, another group of scientists, Allan Maxam and Walter Gilbert, demonstrated their chemical-cleavage method for DNA sequencing. The Maxam-Gilbert method relies on using different chemicals that can cleave the DNA sequence at specific sites, the separation of resulting DNA fragments of variable size using electrophoresis, and deciphering the DNA sequence from the resulting gel bands.
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PCR

PCR

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Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore, it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme

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Related Experiment Video

Updated: Jul 7, 2026

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Published on: May 2, 2025

Polony DNA sequencing.

Gregory J Porreca¹, Jay Shendure, George M Church

¹Harvard Medical School, Boston, Massachusetts, USA.

Current Protocols in Molecular Biology

|February 12, 2008

Summary

This summary is machine-generated.

Polony DNA sequencing offers a cost-effective, high-throughput method for genome resequencing. This technique uses millions of short reads to identify genetic differences compared to a reference genome.

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Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

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DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling

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Area of Science:

Genomics
Molecular Biology
Biotechnology

Background:

Genome resequencing is crucial for understanding genetic variation.
Existing methods can be costly and time-consuming.
There is a need for efficient and accurate high-throughput sequencing technologies.

Purpose of the Study:

To introduce and evaluate Polony DNA sequencing as a method for genome resequencing.
To demonstrate the accuracy and throughput of Polony sequencing.
To enable cost-effective identification of genomic differences.

Main Methods:

Production of mate-paired in vitro shotgun genomic libraries.
Clonal amplification of libraries on microbeads via emulsion PCR.
Sequencing using fluorescent nonamer ligation on a microscope slide.
Alignment of millions of 26-bp reads to a reference genome.

Main Results:

Generation of millions of 26-bp reads per sequencing run.
Successful identification of differences between sequences by read alignment.
Demonstration of Polony sequencing as an inexpensive and accurate approach.

Conclusions:

Polony DNA sequencing is a viable, high-throughput method for genome resequencing.
The technology facilitates cost-effective identification of genetic variations.
This approach significantly contributes to genomic research and applications.