Matthew M. Winkler

DNA-labeled clay: A sensitive new method for tracing particle transport

The behavior of mobile colloids and sediment in most natural environments remains poorly understood, in part because characteristics of existing sediment tracers limit their widespread use. Here we describe the development of a new approach that uses a DNA-labeled montmorillonite clay as a highly sensitive and selective sediment tracer that can potentially characterize sediment and colloid transport in a wide variety of environments, including marine, wetland, ground-water, and atmospheric systems. Characteristics of DNA in natural systems render it unsuitable as an aqueous tracer but admirably suited as a label for tracing particulates. The DNA-labeled-clay approach, using techniques developed from molecular biology, has extremely low detection limits, very specific detection, and a virtually infinite number of tracer signatures. Furthermore, DNA-labeled clay has the same physical characteristics as the particles it is designed to trace, it is environmentally benign, and it can be relatively inexpensively produced and detected. Our initial results show that short (500 base pair) strands of synthetically produced DNA reversibly adsorb to both Na-montmorillonite and powdered silica surfaces via a magnesium bridge. The DNA-montmorillonite surface complexes are stable in calciumbicarbonate spring waters for periods of up to 18 days and only slowly desorb to the aqueous phase, whereas the silica surface complex is stable only in distilled water. Both materials readily release the adsorbed DNA in dilute EDTA solutions for amplification by the polymerase chain reaction (PCR) and quantification. The stability of the DNA-labeled clay complex suggests that this material would be appropriate for use as an extremely sensitive sediment tracer for flow periods of as long as 2 weeks, and possibly longer.

Standards for PCR assays

Publisher Summary The wide versatility of polymerase chain reaction (PCR) has created numerous niches for this technology in research and clinical diagnostic laboratories. In both settings, well-defined DNA and RNA standards, compatible with exquisite sensitivity and specificity of PCR, are needed. Because of their relatively advanced state of development, standards for infectious agents are emphasized in this chapter. PCR-based assays have been formatted as qualitative and quantitative tools. Standards developed for quantitative assays present a special challenge as they must be precisely calibrated. Additionally, independent verification of the amount of very low numbers of nucleic acid templates is restricted by the limited number of corresponding technologies with sensitivities comparable to PCR. The chapter also outlines special problems with standardizing PCR assays. Various types of standards are also discussed including ideal, internal, and external standards. Finally, the discussion concludes by presenting the future needs for standards.