Overview

The Infinium LCG_PGx workflow for the new Global Diversity Array + Enhanced PGx Content (GDA + PGx) includes a new targeted gene amplification (TGA) step. TGA is a PCR amplification for specific pharmacogenomic (PGx) content, which allows for pseudogene disambiguation. The PGx TGA is combined with the Infinium whole genome amplification (WGA) before the Fragment step. The combined sample is processed through the standard LCG Infinium workflow from Fragment on.

The TGA reagents contain a probe that generates a sample-independent TGA positive control fragment. This control fragment binds to the TGA Control Probe on the array when the PGx TGA and WGA are properly executed, recombined, and processed in the Infinium assay. The TGA Control Probe should have a positive signal, observed as Norm R on the Y axis of the SNP graph in GenomeStudio 2.0. The performance expectation for the TGA Control Probe (TGA_Ctrl_5716) is R > 1.

Table 1: GDA+ PGx Product specifications

Evaluate performance of the PGx TGA Control Probe

1. Create the GenomeStudio Genotyping Workspace.
   1. Download and install the current version of GenomeStudio, v2.0.5, from the GenomeStudio Support Resources page
   2. Files required:
      • Intensity data (.idat) files from the scanned BeadChips
      • Manifest: GDA_PGx-8v1-0_20042614_A2.bpm*
      • Cluster file: GDA_PGx-8v1-0_Gentrain_A2.egt*
   3. Create the GenomeStudio workspace as described in the GenomeStudio Genotyping Module v2.0 User Guide
   4. Calculate call rates and evaluate sample performance per specifications in Table 1.
2. Analyze the TGA Control Probe.
   1. Right-click on the Name column in the Full Data Table in GenomeStudio and select Find.
   
   
   

    

   2. In the search window, type “TGA_Ctrl_5716” which is the name of the TGA control probe in the manifest.
   
   
   

    

   3. Examine the SNP Graph to determine if any samples fall below a normalized R (Norm R) of 1. Select the samples with R < 1 to highlight in the SNP Graph and Samples Table for further evaluation.
   
   
   

    

   4. Evaluate the overall performance of samples with a failed TGA control. Determine if there are any other associated run failures by evaluating the data in GenomeStudio, including in the Controls Dashboard.
      If samples do not meet performance expectations for R > 1, exclude them from further analysis, or flag them as having potential inaccuracies in genotype and star allele calls for the PGx content.

*Download product files from the Infinium Global Diversity Array with Enhanced PGx Product Files page.

For additional questions regarding sample and probe performance, contact Illumina Technical Support at techsupport@illumina.com