arget for optimization. Because any natural poliovirus population is heterogeneous, we used in vitro transcribed RNA derived from a plasmid poliovirus clone as a reference set. While TOctober Single Base Extension by Array October Single 6031788 Base Extension by Array parameters were tested throughout the optimization process, with little or no effect on array performance. One important source of noise is experiment-specific and dependent on the relative frequencies of mutations in the population. If a single mutation rises to high levels within a population, it can affect the performance of oligonucleotides assaying nearby positions. Template molecules containing the mutation will hybridize 1201438-56-3 correctly, but mismatches between the template and the MDAP can accurately measure mutation frequency in a controlled population We tested the sensitivity and accuracy of the MDAP platform by measuring the frequency of specific mutants within a defined population. We introduced a set of point mutations into the capsid region of the poliovirus genome and generated the corresponding in vitro transcribed RNA. These mutated genomes were combined with each other and unmodified poliovirus RNA such that each mutant RNA would be present at a frequency of Single Base Extension by Array Monitoring an evolving viral population with MDAP was harvested from each sample upon cytopathic effect and used to infect a fresh monolayer of cells at the same drug concentration. The virus resulting from this second round of infection was termed POctober Single Base Extension by Array Observing the Dynamics of Ribavirin Resistance While drug resistant variants are known to exist in viral populations prior to drug exposure, conventional sequencing-based assays are only able to detect mutants present at a high frequency. The sensitivity and low detection threshold of the MDAP platform offer an opportunity to observe the dynamics of drug resistance at a relatively early stage. We and others have identified a point mutation within the poliovirus Single Base Extension by Array October Single Base Extension by Array MDAP analysis In addition to the MDAP array described here, we have also developed freely available companion software to facilitate rapid and detailed analysis of array results. The open source MDAP software package provides an intuitive graphical user interface, allowing users to load GPR files from 
multiple arrays and generate sample files containing all data relevant to a single sample. Data can be explored through a series of automatically generated graphs, allowing the user to quickly examine data by genome position, fluorescence intensity, and mutation frequency at the level of a single array, sample, or even a reference set of samples. Mutation frequencies, as well as z-scores when a reference set is available, can be exported to text files for further analysis in other programs. The MDAP software package is implemented in python, and can be used on all major computing platforms. The source code is available at http:// derisilab.ucsf.edu/software/MDAP/. A system for accurate measurement of low frequency polymorphisms We devoted considerable effort to optimizing each step of the MDAP protocol. As is the case for many microarray platforms, we found that proper oligonucleotide design is critical. While Discussion Single Base Extension by Array Characterizing the mutant spectrum of an evolving viral population We examined the ability of the MDAP platform to measure chang
