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Related Experiment Video

Updated: Sep 15, 2025

The MultiBac Protein Complex Production Platform at the EMBL

The MultiBac Protein Complex Production Platform at the EMBL

Published on: July 11, 2013

A novel method for generating baculovirus bacmids using EGFP-mediated purification and linearization.

Wujie Su¹, Haoyi Gu¹, Xiaoxia Zhang¹

¹School of Life Sciences, Suzhou Medical College of Soochow University, Soochow University, Suzhou, Jiangsu, 215123, People's Republic of China.

Biotechnology Letters

|July 16, 2025

Summary

This study introduces a novel method for generating baculovirus bacmids using an EGFP reporter for enhanced purification. This technique improves recombinant proportions, particularly for low-titer baculoviruses.

Keywords:

Bac-to-bac system Bacmid construction Baculoviruses BmNPV bacmid Gene deletion Genetic engineering

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Last Updated: Sep 15, 2025

The MultiBac Protein Complex Production Platform at the EMBL

The MultiBac Protein Complex Production Platform at the EMBL

Published on: July 11, 2013

Production and Purification of Baculovirus for Gene Therapy Application

Production and Purification of Baculovirus for Gene Therapy Application

Published on: April 9, 2018

Production of Recombinant PRMT Proteins using the Baculovirus Expression Vector System

Production of Recombinant PRMT Proteins using the Baculovirus Expression Vector System

Published on: July 17, 2021

Area of Science:

Molecular Biology
Virology
Biotechnology

Background:

Baculovirus bacmids are crucial for gene manipulation in insect cells.
Traditional bacmid generation involves homologous recombination or in vitro cloning, which can be inefficient.

Purpose of the Study:

To develop a novel, more efficient method for generating baculovirus bacmids.
To improve the purification and selection process for recombinant baculoviruses.

Main Methods:

Introduction of an EGFP reporter gene into a bacterial DNA cassette.
Generation of an intermediate bacmid with an 11.6 kbp cassette containing EGFP.
Purification of recombinants in cultured cells using EGFP as a reporter.
Linearization of the intermediate bacmid and co-transfection with a PCR-amplified cassette into BmN cells for homologous recombination.
Transformation of DH10B competent cells to obtain the final bacmid.

Main Results:

The novel method successfully generated baculovirus bacmids.
The use of EGFP facilitated purification, increasing the proportion of recombinants.
The method is effective even for baculoviruses with low virus production titers.

Conclusions:

The developed method offers a significant improvement for baculovirus bacmid generation.
This approach enhances efficiency and is particularly beneficial for challenging baculovirus systems.
The EGFP-mediated purification strategy increases the overall success rate of bacmid construction.