- MoChlo: A versatile modular cloning toolbox for chloroplast biotechnology
- Golden Mutagenesis: An efficient multi-site saturation mutagenesis approach by Golden Gate cloning with automated primer design
- CyanoGate: A Golden Gate modular cloning suite for engineering cyanobacteria based on the plant MoClo syntax
- Birth of a photosynthetic chassis: a MoClo toolkit enabling synthetic biology in the microalga Chlamydomonas reinhardtii
- Loop Assembly: a simple and open system for recursive fabrication of DNA circuits
- AQUA Cloning: A Versatile and Simple Enzyme-Free Cloning Approach
- GoldenBraid 2.0: A Comprehensive DNA Assembly Framework for Plant Synthetic Biology
- A Golden Gate Modular Cloning Toolbox for Plants
- A versatile and robust Agrobacterium‐based gene stacking system generates high‐quality transgenic Arabidopsis plants
- A robust gene-stacking method utilizing yeast assembly for plant synthetic biology
AQUA Cloning: A Versatile and Simple Enzyme-Free Cloning Approach
Assembly cloning is increasingly replacing conventional restriction enzyme and DNA-ligase-dependent cloning methods for reasons of efficiency and performance. Here, we describe AQUA (advanced quick assembly), a simple and versatile seamless assembly cloning approach. We demonstrate the applicability and versatility of AQUA Cloning in selected proof-of-principle applications including targeted insertion-, deletion- and site-directed point-mutagenesis, and combinatorial cloning. Furthermore, we show the one pot de novo assembly of multiple DNA fragments into a single circular plasmid encoding a complex light- and chemically-regulated Boolean A NIMPLY B logic operation. AQUA Cloning harnesses intrinsic in vivo processing of linear DNA fragments with short regions of homology of 16 to 32 bp mediated by Escherichia coli. It does not require any kits, enzymes or preparations of reagents and is the simplest assembly cloning protocol to date.
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