Assay Processing

Second Genome's analysis can measure the microbial composition of any sample type, including human and mouse sources, fecal samples, soil, air, and frozen water filters. We accept both isolated DNA and frozen water filters. For isolated DNA, we request a minimum of 200 ng/10 µl DNA per assay type.

We also offer a variety of solutions for biological specimen collection that enable reproducible, high quality DNA isolation. These following includes protocols and kits  for sample collection and shipping of human feces, animal feces, swabs, frozen water filters, and soil.

The entire 16S rRNA gene is amplified from DNA extracts. For each sample, amplified products are purified and quantified. PhyloChip Control Mix^™ is added for batch and quality control as well as for scaling and normalization for quantification. The combined mixture for each sample is then fragmented, biotin labeled, and hybridized to the PhyloChip array, version G3. PhyloChip arrays are washed, stained, and scanned.

Quality Control and Performance

Samples are processed by experienced professionals in a Good Laboratory Practices (GLP) compliant service laboratory running Quality Management Systems for sample and data tracking. The laboratory implements detailed SOPs, equipment and process validation, training, audits and document control measures. QC and QA metrics are maintained for all sample handling, processing and storage procedures.

The PhyloChip assay demonstrates high reproducibility from array-to-array and from run-to-run. Measurements are also highly quantitative and reliable, even for low abundance organisms.  To assess the performance of the third generation PhyloChip assay, twenty-six 16S rDNA mixtures from different species were prepared as mock communities using a semi-randomized Latin square structure. Overall, the variations were minor producing a mean CV = 0.097. Additionally, a significant correlation was found between the concentrations of each gene in the Latin Square and the corresponding hybridization score generating an average correlation coefficient, r = 0.941.