Method for efficient soluble expression and purification of recombinant hyperactive Tn5 transposase.
Tongtong Xu1, Minqin Xiao1, Lin Yu1
1State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University and Collaborative Innovation Center of Biotherapy, 610041, Chengdu, China.
Protein Expression and Purification
|March 15, 2021
Summary
We developed a cost-effective method for preparing next-generation sequencing (NGS) libraries using a modified Tn5 transposase. This improved purification strategy ensures high activity and low contamination, making NGS library construction more accessible.
Area of Science:
- Molecular Biology
- Genomics
- Biotechnology
Background:
- Efficient library preparation is crucial for next-generation sequencing (NGS).
- Tn5 transposase facilitates robust and efficient tagmentation-based library construction.
- Existing methods can be costly and complex.
Purpose of the Study:
- To develop a simplified and reliable expression and purification strategy for Tn5 transposase.
- To reduce the costs associated with Tn5-based NGS library construction.
- To enhance the utility of Tn5 transposase for novel NGS applications.
Main Methods:
- Fusion of Tn5 transposase to the GB1 domain and a 10× His tag for expression and purification.
- Purification of recombinant Tn5 transposase.
- Assay of DNA tagmentation activity and nucleic acid contamination levels.
Main Results:
- Successfully expressed and purified recombinant Tn5 transposase with high activity.
- Achieved ultra-low nucleic acid contamination in the purified enzyme.
- Demonstrated a cost-effective approach to Tn5-based NGS library preparation.
Conclusions:
- The developed strategy provides a simple, reliable, and cost-effective method for Tn5 transposase purification.
- This approach lowers the barrier to entry for Tn5-based NGS library construction.
- The findings support the advancement of new NGS methodologies.
