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DNA Isolation

DNA Isolation

DNA isolation protocols can be fast and straightforward or complex and time-consuming depending on the type and quality of DNA required for further processing. For example, plasmid DNA extraction is a bit more complicated than genomic DNA extraction because of the need for an appropriate lysis method to separate plasmid DNA from gDNA during isolation. However, for specific applications, such as long-range DNA sequencing that require a good yield of high- quality DNA samples, we need to follow...

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Updated: Jun 5, 2025

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Published on: December 1, 2017

RH2Fusion: A universal tool for precise DNA fragment assembly.

Benchao Liu¹, Junru Zhao¹, Hui Chen¹

¹State Key Laboratory of Cellular Stress Biology, Innovation Centre for Cell Signalling Network, Engineering Research Centre of Molecular Diagnostics of the Ministry of Education, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China.

International Journal of Biological Macromolecules

|December 15, 2024

Summary

This summary is machine-generated.

RNase HII Fusion (RH2Fusion) enables scarless DNA assembly with user-defined sticky ends and multiple site-directed mutagenesis. This novel system utilizes specific enzyme activity for efficient DNA fragment joining and modification, advancing molecular biology research.

Keywords:

DNA assembly Molecular cloning RH2Fusion RNase HII Ribonuclease YgdG

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Rapid Assembly of Multi-Gene Constructs using Modular Golden Gate Cloning

Rapid Assembly of Multi-Gene Constructs using Modular Golden Gate Cloning

Published on: February 5, 2021

Area of Science:

Molecular Biology
Synthetic Biology
Genetic Engineering

Background:

Restriction enzyme (RE)-mediated cleavage is the standard for DNA assembly but has limitations.
Generating sticky ends for DNA assembly is crucial for molecular biology techniques.

Purpose of the Study:

To introduce RNase HII Fusion (RH2Fusion), a novel system for scarless DNA assembly.
To enable user-defined sticky ends and simultaneous site-directed mutagenesis (SDM).

Main Methods:

RH2Fusion utilizes RNase HII and Escherichia coli Exonuclease IX (YgdG) for DNA fragment processing.
In vivo, ribonucleotide-modified DNA fragments form 3' overhangs after RNase HII treatment for recombination.
In vitro, RNase HII and YgdG generate overhangs processed by YgdG and T4 DNA ligase.

Main Results:

RH2Fusion allows scarless assembly of multiple DNA fragments.
Simultaneous site-directed mutagenesis (SDM) at multiple sites is achievable.
Escherichia coli Exonuclease IX (YgdG) demonstrates specific ribonuclease activity.

Conclusions:

RH2Fusion provides a rapid, effective, and versatile alternative for DNA assembly.
The system enhances DNA manipulation capabilities in synthetic biology and genetic engineering.
This tool advances molecular biology research by improving DNA assembly and modification techniques.