Quality assurance samples submitted from the NCSBI as part of a contract with TBTG to outsource DNA Database samples showed unexpected discrepancies for the locus D16S539 when all other loci yielded identical results. Discrepancies observed included allele drop out and an imbalance in sister alleles with samples returned from TBTG. This led to a comprehensive review of the technical procedures used between the two laboratories to determine the cause of the discrepancies noted for the locus D16S539, since both laboratories were using the PowerPlex® 1.1 typing kit from the Promega Corporation. The NCSBI and the TBTG utilize different extraction methods (organic extraction vs. FTA) and amplification conditions (AmpliTaq® vs AmpliTaq Gold®), respectively, so the exact cause of discrepancy observed was not immediately apparent. Experiments at the NCSBI associated the observed allele drop out and the imbalance of the sister alleles with the use of AmpliTaq Gold® and a hot start procedure. Sequencing data revealed that a point mutation resides on the D16S539 primer-binding site that reaches polymorphic levels in African-American populations. This led to the development of a degenerate primer by the Promega Corporation to detect “missing” alleles when AmpliTaq Gold® is used. The degenerate primer was then thoroughly tested to show its efficacy in detecting the “true” D16S539 profile when used.

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Small columns of precisely deactivated silicic acid (3 g, 3.3% water) were investigated for separating hexachlorobenzene, polychlorinated biphenyls (PCBs), DDE, DDT, and cis- and trans-chlordane from toxaphene. The chlorinated hydrocarbons were eluted from the column in 3 fractions: Fraction 1 (50 ml petroleum ether) contained HCB, PCBs, DDE, and 10-30% o,p'-DDT. If desired, HCB could be separated from PCBs by further dividing fraction 1 into 2 portions, the first 13 ml containing HCB and the next 37 ml containing PCBs and DDE. Fraction 2 (80 ml petroleum ether) contained the chlordanes, p,p'-DDT, and 70–90% o,p'-DDT. Dieldrin and p,p'-DDD were eluted in a third (15 ml dichloromethane) fraction. The percentages of toxaphene eluted in the 3 fractions were approximately 10, 30, and 60%, respectively. Most of the toxaphene components interfering with the analysis of p,p'- DDT were eluted in the third fraction. This procedure has been very helpful in separating and identifying many of the high molecular weight chlorinated hydrocarbons in ambient air.