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Related Experiment Video

Updated: Aug 21, 2025

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

A new combinatorial megaplasmid library assembly method designed to screen for minimal pathways by using SCRaMbLE.

Wei Sheng Yap^1,2, Guillaume Thibault^1,3,4

¹School of Biological Sciences, Nanyang Technological University, Singapore, 637551.

Micropublication Biology

|November 17, 2022

Summary

This summary is machine-generated.

Researchers developed a cost-effective method for assembling human gene pathways in yeast. This high-throughput technique creates a yeast megaplasmid library for screening minimal functional pathways, aiding protein production and directed evolution.

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Published on: October 3, 2018

Area of Science:

Synthetic biology
Molecular biology
Biotechnology

Background:

Yeast is widely used for expressing human proteins to improve production.
Establishing functional human pathways in yeast remains a significant challenge.
High-throughput screening methods are needed for pathway discovery.

Purpose of the Study:

To develop a simple, economical, and high-throughput method for assembling human functional pathways in yeast.
To create a yeast megaplasmid library from human cDNA for screening minimal pathways.
To facilitate directed evolution experiments and adaptation in other organisms.

Main Methods:

Utilized Golden Gate assembly for gene construction.
Introduced artificial promoters and symmetric loxP sites into yeast megaplasmids.
Employed streptavidin-bead-based purification for isolating assembled DNA.

Main Results:

Successfully created a high molecular weight, randomly assembled human cDNA megaplasmid library in yeast.
The method is simple, economical, and suitable for high-throughput screening.
The library is poised for screening minimal human functional pathways.

Conclusions:

The developed gene assembly method provides a powerful tool for constructing complex genetic libraries in yeast.
This approach can accelerate the discovery of minimal functional pathways for various applications.
The methodology is adaptable for use in other model organisms for synthetic biology applications.