A PCR-based method for isolation of genomic DNA flanking a known DNA sequence

We describe a simple PCR-based method for the isolation of genomic DNA that lies adjacent to a known DNA sequence. The method is based on the directional cloning of digested genomic DNA into the multiple cloning site of a pUC-based plasmid to generate a limited genomic library. The library is plated onto a number of selective LA plates which are incubated overnight, and recombinant plasmid DNA is then isolated from resistant colonies pooled from each plate. PCR amplification is performed on the pooled recombinant plasmid DNAs using primers specific for the pUC vector and the known genomic sequence. The combination of efficient directional cloning and bacterial transformation gives relative enrichment for the genomic sequence of interest and generates a simple DNA template, enabling easy amplification by PCR

Rapid localization of proteins in chromatographic eluates

Rapid localization of proteins in chromatographic eluate

The separation of malic dehydrogenase isoenzymes of Neurospora crassa by polyacrylamide gel electrophoresis

The separation of malic dehydrogenase isoenzymes of Neurospora crassa by polyacrylamide gel electrophoresi

In-vehicle nitrogen dioxide concentrations in road tunnels

There is a lack of knowledge regarding in-vehicle concentrations of nitrogen dioxide (NO) during transit through road tunnels in urban environments. Furthermore, previous studies have tended to involve a single vehicle and the range of in-vehicle NO concentrations that vehicle occupants may be exposed to is not well defined. This study describes simultaneous measurements of in-vehicle and outside-vehicle NO concentrations on a route through Sydney, Australia that included several major tunnels, minor tunnels and busy surface roads. Tests were conducted on nine passenger vehicles to assess how vehicle characteristics and ventilation settings affected in-vehicle NO concentrations and the in-vehicle-to-outside vehicle (I/O) concentration ratio. NO was measured directly using a cavity attenuated phase shift (CAPS) technique that gave a high temporal and spatial resolution. In the major tunnels, transit-average in-vehicle NO concentrations were lower than outside-vehicle concentrations for all vehicles with cabin air recirculation either on or off. However, markedly lower I/O ratios were obtained with recirculation on (0.08–0.36), suggesting that vehicle occupants can significantly lower their exposure to NO in tunnels by switching recirculation on. The highest mean I/O ratios for NO were measured in older vehicles (0.35–0.36), which is attributed to older vehicles having higher air exchange rates. The results from this study can be used to inform the design and operation of future road tunnels and modelling of personal exposure to NO