Polynucleotide contamination

Many benefits will be realized by incorporating these polynucleotide arrays into an integrated analysis system. Decreased assay cost and contamination, along with improved throughput and reproducibility can be achieved by integrating sample preparation processes with the polynucleotide arrays these systems under development at Affymetrix [20-21]. 

As for any analytical technique, a major problem lies in the purity of the polynucleotide sample, although nucleoprotein contamination is not prohibitive to the recording of FAB spectra. Furthermore, it must be noted that branched polynucleotides cannot be analyzed with our programs, exception made of the fragmentary information obtained from all ends. 

If ferrous ions are present during assays of DNA and RNA polymerases in which radioactive newly synthesized polynucleotides are precipitated on to filter paper discs in trichloroacetic acid, spuriously high results are obtained, apparently due to acid-insoluble complexes of Fe with substrate nucleotides precipitating on the paper. Clearly ferrous ions should be avoided in assays of this type. The complex Al. ATP is a potent inhibitor of hexokinase, and neutron activation analysis of many commercial samples of ATP has shown that Al is ubiquitous, and the most common metal contaminant It is best removed by passing the ATP preparation over a cellulose polyphosphate column at pH 5. Chelates of ATP with divalent metal ions have been separated from non-chelated ATP using reverse-phase h.p.l.c. ... 

Restriction enzyme activities can be assayed by various methods including gel visualization assay, polynucleotide kinase exchange assay (36), and exonuclease-coupled assay (37). By far the most common of all assay methods is the visualization by EtBr staining of the digestion products separated by electrophoresis in an agarose gel (30,38). This method is straightforward and enables the detection of contaminating activities of endo- and exonucleases. 

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