Application of environmental DNA resources to create useful DNA polymerases with different properties

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

DNA polymerases use deoxynucleotide triphosphates to synthesize new DNA strands according to the template DNA during DNA replication and repair, and are essential to maintain genome integrity in DNA metabolism. In addition, these enzymes are widely used for genetic engineering techniques, including dideoxy-sequencing, PCR, DNA labeling, mutagenesis, and other in vitro gene manipulations. Thermostable DNA polymerases are especially useful for PCR and cycle-sequencing. We describe in this chapter a powerful strategy to use environmental DNA as a genetic resource to create useful DNA polymerases. The region corresponding to the active center of the DNA polymerizing reaction in the structural gene of well known DNA polymerases, such as Pfu DNA polymerase and Taq DNA polymerase, can be substituted with gene fragments amplified by PCR from DNAs within soil samples from various world-wide locations. The constructed chimeric pol genes can be expressed in E. coli, and the produced chimeric enzymes, possessing DNA polymerase activities with different properties, can be evaluated in terms of their processivity, fidelity, and efficiency of primer usage, to select valuable DNA polymerases for genetic engineering techniques.

Original languageEnglish
Title of host publicationMicroorganisms in Sustainable Agriculture and Biotechnology
PublisherSpringer Netherlands
Pages663-678
Number of pages16
Volume9789400722149
ISBN (Electronic)9789400722149
ISBN (Print)9400722133, 9789400722132
DOIs
Publication statusPublished - Oct 1 2012

All Science Journal Classification (ASJC) codes

  • Immunology and Microbiology(all)

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  • Cite this

    Ishino, S., & Ishino, Y. (2012). Application of environmental DNA resources to create useful DNA polymerases with different properties. In Microorganisms in Sustainable Agriculture and Biotechnology (Vol. 9789400722149, pp. 663-678). Springer Netherlands. https://doi.org/10.1007/978-94-007-2214-9_29