Affinity purification of plasmid DNA by temperature-triggered precipitation

作者：Kostal J; Mulchandani A; Chen W^*

来源：Biotechnology and Bioengineering, 2004, 85(3): 293-297.

DOI：10.1002/bit.10890

• DNA cloning
• plasmid binding
• plasmid purification
• affinity precipitation
• elastin-like proteins (ELPs)

This report describes a new plasmid DNA purification method, which takes advantage of the DNA-binding affinity and specificity of the bacterial metalloregulatory protein MerR, and of the temperature responsiveness of elastin-like proteins (ELPs). Upon increasing the temperature, ELP undergoes a reversible phase transition from water-soluble forms into aggregates, and this property was exploited for the precipitation of plasmid DNA containing the MerR recognition sequence by a simple temperature trigger. In one purification step, plasmid DNA was purified from E. coli cell lysates to a better purity than that prepared by a standard alkaline purification method, with no contaminating chromosomal DNA and cellular proteins. This protein-based approach, in combination with the reversible phase transition feature of ELP, makes the outlined method a promising candidate for large-scale purification of plasmid DNA for sensitive applications such as nonviral gene therapy or DNA vaccines.

• 出版日期2004-2-5