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Cleavage of bacteriophage M13 DNA by Haemophilus influenzae endonuclease-R

C M Van den Hondel1, J G Schoenmakers

  • 1Laboratory of Molecular Biology, University of Nijmegen, Nijmegen, The Netherlands.

Molecular Biology Reports
|November 8, 2013
PubMed

Insights

The restriction enzyme endonuclease-R from Haemophilus influenzae precisely cuts bacteriophage M13 DNA at a single site. This generates linear DNA molecules, demonstrating the enzyme's high specificity in DNA cleavage.

Area of Science:

  • Molecular Biology
  • Enzymology
  • Genetics

Background:

  • Restriction enzymes are crucial tools in molecular biology for DNA manipulation.
  • Haemophilus influenzae endonuclease-R is a specific type of restriction enzyme.
  • Bacteriophage M13 DNA is a commonly studied model system in genetic research.

Purpose of the Study:

  • To characterize the cleavage site specificity of endonuclease-R from Haemophilus influenzae.
  • To investigate the enzymatic activity of endonuclease-R on bacteriophage M13 replicative form DNA.

Main Methods:

  • Enzymatic digestion of double-stranded replicative form DNA of bacteriophage M13 using endonuclease-R.
  • Analysis of DNA fragment lengths and structural integrity post-digestion.

Main Results:

  • Endonuclease-R exhibits a single, specific cleavage site on bacteriophage M13 replicative form DNA.
  • The enzyme produces full-length linear M13-DNA molecules (RF-III).
  • Generated RF-III molecules are incapable of re-circularization, confirming site specificity.

Conclusions:

  • Endonuclease-R demonstrates high sequence specificity in DNA cleavage.
  • This enzyme is a valuable tool for generating linear DNA fragments from circular M13 DNA.

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