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Stepwise relaxation of supercoiled SV40 DNA
Cold Spring Harbor Symposia on Quantitative Biology
|January 1, 1975
Summary
Agarose gel electrophoresis can detect subtle changes in supercoiled DNA structure. This method helped isolate a protein that removes superhelical turns, revealing SV40 DNA has 20-24 turns.
Area of Science:
- Molecular Biology
- Biochemistry
Background:
- Supercoiled DNA plays a crucial role in cellular processes.
- Understanding DNA tertiary structure is essential for molecular biology research.
Purpose of the Study:
- To investigate the utility of agarose gel electrophoresis for analyzing supercoiled DNA tertiary structure.
- To identify and characterize proteins that modify superhelical turns in DNA.
Main Methods:
- Agarose gel electrophoresis was employed to examine supercoiled Simian Virus 40 (SV40) DNA.
- A protein purification assay was developed using the DNA relaxing activity.
Main Results:
- Agarose gel electrophoresis demonstrated sensitivity to detect differences as small as one superhelical turn.
- A DNA relaxing protein was purified from human tissue culture cells.
- The purified protein gradually removed superhelical turns from supercoiled DNA.
- It was deduced that native SV40 DNA possesses a minimum of 20 physical superhelical turns, with an upper limit not exceeding 24.
Conclusions:
- Agarose gel electrophoresis is a viable method for studying supercoiled DNA tertiary structure.
- The purified protein is a key factor in regulating DNA superhelicity.
- The determined number of superhelical turns in SV40 DNA aligns with previous findings.