COMPARATIVE ANALYSIS OF METHODS FOR THE PURIFICATION OF DNA FROM LOW-BIOMASS SAMPLES BASED ON TOTAL YIELD AND CONSERVED MICROBIAL DIVERSITY

Despite advances in the specificity and sensitivity of molecular biological technologies, the efficient recovery of DNA from low-biomass samples remains extremely challenging. Optimal methods to purify biomolecules from such environments should (1) achieve the greatest total yield and (2) reflect the true microbial diversity of the sample. These attributes were assessed from five DNA purification regimes: a standard-manual procedure, MoBio Ultraclean and Promega Wizard kits, and an automated Axcyte AutoLyser method with and without bead-beating agitation. A homogenous mixture of known concentrations of nine distinct bacterial lineages isolated from low-biomass environments was prepared and suitable aliquots of subsamples were processed in parallel. DNA products from each of these methods were then subjected to polymerase chain reaction (PCR), quantitative PCR and 16S rRNA clone-library analysis. The Axcyte AutoLyser outperformed all other purification regimes examined. This automated method consistently both yielded the highest concentration of PCR-amplifiable DNA, and reported species composition most consistent with the starting solution.

PRACTICAL APPLICATIONS

This communication carefully examines the effectiveness of common DNA purification regimes as well as an automated method. Comparative analyses convincingly demonstrate that the different methods not only result in different recovery of genomic DNA, but more importantly, different estimations of microbial diversity in the sample. This report will hopefully inspire investigators from various industries (pharmaceutical, ecological, medical, semiconductor, etc.) who find themselves in the initial phases of large-scale studies to devote a significant effort into optimizing sample extraction protocols to achieve the most accurate information.     

Plasmid analysis of cadmium- and penicillin-resistant strains of staphylococci isolated in maternity hospitals]

The plasmid analysis of a collection of the staphylococci isolated was made in the obstetric hospitals of Nizhny Novgorod and Arzamas in 1986-1989. It revealed the presence of large complex plasmids with a molecular weight of 22 MD carrying the determinants of resistance to penicillins and cadmium and mercury ions in polyresistant strains. Conjugation transfer of the plasmid even between the staphylococci of different species was performed under experimental conditions. It was suggested that occurrence of the strains carrying the complex cadmium-penicillinase plasmid was due to violation of the sanitary or ecological requirements.     

Differences in morphology and colour pattern of shiny cowbird (Molothrus bonariensis) eggs found in nests of two hosts

Genetic differentiation among shiny cowbird (Molothrus bonariensis) females that use different hosts indicates that in this brood parasite, host use is not random at an individual level. We tested whether there exist differences in morphology and coloration between eggs of shiny cowbirds laid in the nests of two different hosts, the chalk‐browed mockingbird (Mimus saturninus) and the house wren (Troglodytes aedon). We took morphometric measures of shiny cowbird eggs found in nests of mockingbirds and wrens and analysed their coloration using digital photography and reflectance spectrometry. We found that shiny cowbird eggs found in mockingbird nests were wider and more asymmetric than those found in wren nests. In addition, cowbird eggs coming from mockingbird nests were brighter and had higher relative red reflectance than those coming from wren nests. Our results show that shiny cowbird eggs laid in nests of two different hosts vary in shape and background colour, but not in spotting pattern. © 2011 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 102 , 838–845.