Large Scale Preparation of Positively Supercoiled DNA Using the Archaeal Histone HMf | Zendy

William A. LaMarr | Zendy; Kathleen Sandman | Zendy; John N. Reeve | Zendy; Peter C. Dedon | Zendy

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Large Scale Preparation of Positively Supercoiled DNA Using the Archaeal Histone HMf

Author(s) -

William A. LaMarr,

Kathleen Sandman,

John N. Reeve,

Peter C. Dedon

Publication year - 1997

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/25.8.1660

Subject(s) - dna supercoil , biology , puc19 , dna , recombinant dna , histone , plasmid , biophysics , xylanase , linking number , biochemistry , enzyme , gene , dna replication

A technique to prepare relatively large quantities (>/=100 microg) of highly positively supercoiled DNA is reported. This uses a recombinant archaeal histone (rHMfB) to introduce toroidal supercoils, and an inexpensive chicken blood extract to relax unrestrained superhelical tension. Preparation of positively supercoiled pUC19 DNA molecules, >50% of which have linking number changes ranging from+8 to+17, is demonstrated. Advantages include the high degree of positive supercoiling that can be achieved, control over the extent of supercoiling, easy production of relatively large quantities of supercoiled DNA, and low cost.

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