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Purified Single-Stranded M13 DNA for Automated Sequencing

that obtained from the SGB method (Figure 1). For 3 ml of phage lysate, the StrataPrep method yielded 7.5 g of DNA at a concentration of 25 ng/l, as judged by gel electrophoresis.

Figure 1

Quality Automated Sequencing

Equal volumes of StrataPrep- and SGB-purified Phagescript DNA were subjected to fluorescent dye primer automated sequencing by an independent sequencing service (Sequetech Corp.; Figure 2). Signal intensity was more than twice as high for the StrataPrep-purified template (Figure 2, Panel A) compared to the SGB-purified template (Figure 2, Panel B). Sequence read length was nearly 700 bases for both templates. There were three and four sequence ambiguities scored for the StrataPrep- and SGB-purifed templates, respectively, within the first 20 nucleotides. Sequence ambiguities also occurred at nucleotides 553 and 638 for the StrataPrep-purified template and at nucleotide 44 for the SGB-purified template.

Figure 2

Conclusion

The StrataPrep M13 DNA purification kit contains a fast, easy, and reliable method for M13 purification. Recover single-stranded DNA from phage lysate more efficiently using the StrataPrep method than with the leading commercial silica-gel-based method, and realize high-quality automated fluorescent sequencing results as well. Additionally, the kit can be used to purify single-stranded DNA from rescued phagemid clones.

REFERENCES
  1. Richards, S., et al. (1994) In Automated DNA Sequencing and Analysis (Eds. Adams, M.D., et al.) pp. 191-198. Academic Press, San Diego.
  2. Wilson, R.K. and Mardis, E.R. (1997) In Genome Analysis: A Laboratory Manual (Eds. Birren, B., et al.) Vol.
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