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Purified Single-Stranded M13 DNA for Automated Sequencing


A streamlined and reliable method to obtain high-quality single-stranded DNA

Jeff Braman Scott Basehore
Stratagene

The StrataPrep M13 DNA purification kit allows easy, quick, and reliable purification of bacteriophage single-stranded DNA. The yield of single-stranded DNA recovered from phage lysate exceeds the yield obtained with the leading commercial silica-gel-based (SGB) M13 DNA purification method. The resulting high-purity DNA is ideal for automated sequencing applications.

M13 DNA is a preferred template for many sequencing projects.1-5 For many years, the standard laboratory protocol used to purify M13 DNA relied upon polyethylene glycol (PEG)-NaCl precipitation of phage particles, phenol-chloroform extraction to remove phage coat proteins, and ethanol precipitation to concentrate the DNA.6 This method is time-consuming, employs hazardous chemicals, and is difficult to apply to more than a few samples.

However, with the StrataPrep M13 DNA purification kit, isolating high-purity M13 DNA is possible without these cumbersome steps because the procedure is streamlined: Collect M13 phage particles on the unique glass fiber matrix contained in a StrataPrep spin cup;7 add a chaotropic salt solution to the spin cup so the phage coat protein denatures and binds the liberated single-stranded DNA to the glass matrix; centrifuge-wash the phage DNA to remove contaminating protein and salt; and elute the DNA from the spin cup with a low ionic strength buffer. The concentration of the highly purified DNA that results is ideal for automated sequencing applications.

High Yield Recovery of Single-Stranded DNA

The yield of single-stranded DNA from StrataPrep M13 method of Phagescript lysate (a modified M13 bacteriophage8), was at least twice
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