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Solar UV Radiation

Solar UV Radiation. UVC <290 nm Absorbed by ozone UVB 290 – 320 nm ~0.5 % total energy UVA 320 – 400 nm 6% total energy. UV Spectrum of DNA. Absorbance. 300. 250. Wavelength (nm). T<>T. T<>C. C<>C. Pyrimidine Photodimers. Bipyrimidine (6-4) Photoadducts.

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Solar UV Radiation

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  1. Solar UV Radiation • UVC <290 nm Absorbed by ozone • UVB 290 – 320 nm ~0.5 % total energy • UVA 320 – 400 nm 6% total energy

  2. UV Spectrum of DNA Absorbance 300 250 Wavelength (nm)

  3. T<>T T<>C C<>C Pyrimidine Photodimers

  4. Bipyrimidine (6-4) Photoadducts

  5. DNA Photooxidation Mechanisms Oxidation at GG(A) doublet P G Oxidation hν Hole Transfer DNA Type I Type II P-.+ DNA+. P* 1O2 singlet oxygen O2 DNA

  6. Method of irradiation and visualization of DNA damage UV irradiation 32P-DNA + PS Upturned Eppendorf lid EtOH precipitation  Piperidine Electrophoresis Autoradiography

  7. Single-stranded DNA • Singlet oxygen • UV dose & [SP] dependent Uniform G-specific attack

  8. Quenching in ssDNA 1SP + UVB 2 & 3Quenched by sodium azide & DABCO 4 & 5Mannitol & SOD have no effect

  9. H2O D2O Singlet oxygen damage in ss DNA increases in D2O

  10. Duplex DNA Specificity for 5’-G of 5’-GG-3’ doublets 5’-G of 5’-GA-3’ doublets

  11. Conclusions • Single-stranded DNA - Type II mechanism • singlet oxygen • Duplex DNA - Type I mechanism • electron transfer - DNA to excited sensitizer • hole migration

  12. MALDI-TOF Analysis of DNA • Requires picomole amounts of sample • Mass accuracy to ~0.1% • Generate +ve and –ve ion spectra • UV-absorbing matrix for <100 base pairs • Applications in DNA sequencing

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