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Mismatch Repair

Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the new strand and replacing the erroneous bases with correct nucleotides. Mismatch repair is coordinated by many proteins in both prokaryotes and eukaryotes.
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...

Mismatch Repair

Mismatch Repair

Fixing Double-strand Breaks

Fixing Double-strand Breaks

The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...

Fixing Double-strand Breaks

Fixing Double-strand Breaks

The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...

Homologous Recombination

Homologous Recombination

The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...

Proofreading

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: Jun 3, 2026

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Published on: December 1, 2017

From fragmented workflows to integrated pipelines: Bridging enzymatic DNA synthesis, assembly, and MutS-based error

Xiaohang Wang¹, Xinran Zhang², Wenfei Yu³

¹University of Jinan, Jinan, Shandong, China; Single-Cell Center, Key Laboratory of Photoelectric Conversion and Utilization of Solar Energy, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101, China.

Biotechnology Advances

|June 1, 2026

Summary

Integrating DNA synthesis, assembly, and error correction is key for scalable gene construction. A continuous, automated workflow using enzymatic DNA synthesis and MutS-based error correction enhances fidelity and throughput.

Keywords:

Automation Enzymatic DNA synthesis Error correction Fragment assembly High-fidelity DNA synthesis MutS-based mismatch depletion Synthetic biology Workflow integration

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Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis

Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis

Published on: June 19, 2018

Related Experiment Videos

Last Updated: Jun 3, 2026

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Published on: December 1, 2017

Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis

Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis

Published on: June 19, 2018

Area of Science:

Synthetic Biology
Molecular Biology
Biotechnology

Background:

Gene construction at the gene-to-pathway scale is crucial for synthetic biology.
Current methods for oligonucleotide synthesis, fragment assembly, and error correction are fragmented and disconnected.
This fragmentation limits the fidelity, throughput, and scalability of DNA sequence construction, especially for large sequences.

Purpose of the Study:

To review the challenges in current gene construction pipelines.
To highlight the importance of integrating synthesis, assembly, and error correction.
To propose a new, integrated, and automated workflow for high-fidelity DNA construction.

Main Methods:

Tracing error propagation from oligonucleotide synthesis through hierarchical assembly.
Comparing different DNA error correction strategies.
Evaluating the potential of enzymatic DNA synthesis for pipeline integration.

Main Results:

Errors in oligonucleotide synthesis significantly impact downstream assembly.
MutS-based enzymatic mismatch depletion demonstrates superior error correction efficiency and workflow compatibility.
Enzymatic DNA synthesis enables aqueous, downstream-compatible processes.

Conclusions:

An integrated pipeline, not just sequential automation, is necessary for scalable gene construction.
Combining enzymatic DNA synthesis with MutS-based error correction offers a path to a continuous, automated workflow.
This integrated approach promises high-fidelity, high-throughput, and scalable DNA sequence construction.