The mouse has become the most widely used mammalian model for basic biological and disease discovery research. The explosion in mouse genomic sequence availability in recent years has enabled the development of a whole genome microarray with the potential to revolutionize fundamental research. Agilent has taken extra measures to design this microarray to represent the best possible view of the mouse genome as we know it today.
| Feature Layout |
|
Agilent's SurePrint 60-mer in situ manufacturing process enables true flexibility in microarray layout and format for generation of the Whole Mouse Genome microarray. |
|
| 60-mer Probe Selection with a GeneBin
A GeneBin is defined herein as a grouping of transcript sequences that share some sequence identity and overlap on the genome.
|
|
(A) Demonstration of selection of consensus regions and candidate probes to represent transcripts within a GeneBin. The transcript sequences within a GeneBin (shown at top) are used to determine the location of consensus regions (shown in the middle). In this example, three different consensus regions were chosen and probes selected from those consensus regions (shown at bottom). |
|
(B) Clustering of data demonstrates that probes for consensus regions 2 and 3 exhibit similar behavior, while probes for consensus region 1 exhibit a different behavior, thus representing, in this case, a real alternative 3' end of transcript A. In the above figures, probes are colored based on the supercluster to which they belong. Probes colored gray did not cluster with other probes. Although three consensus regions were designed in this example, only two probes were selected for this GeneBin (shown by the star), one for each supercluster. |
|
High Reproducablity
Defining the level of reproducibility in a microarray system provides an understanding of the consistency of the system in measuring differential expression.
|
|
Inter-array reproducability measures the performance consistency across different microarrays. It was determined using eight replicate self vs. self experiments. Data points1 represent standard deviations (SDs) of the log ratios for each probe across eight replicate hybridizations. The Median SD across all data points < 0.032. |
|
Intra-array reproducability measures the performance consistency within a microarray. It was determined using four replicate self vs. self experiments, each represented by a different color. Data points1 represent standard deviations of the log ratios across replicate probes within the same microarray (each microarray contains ten replicates for each of forty unique probes). The Median SD across 10 replicate probes ranges from 0.017 to 0.025. |
|
| High Quality Data |
|
System noise, a measure of the non-systematic variability in the labeling, hybridization and scanning processes, provides an understanding of the sensitivity and precision of the microarray system in measuring differential expression. Low system noise leads to more consistent data and enhanced sensitivity as demonstrated by the results of a self vs. self hybridization. Data points are color-coded based on whether the log ratio values are determined to be significantly different than 0 (P < 0.01) by the Agilent's Feature Extraction software (v. 7.5). 99.86% of all probes examined do not show differential expression (shown in yellow). (Yellow - Unchanged; Red - Up-regulated; Green - Down-regulated). |
|
| Unparalleled Sensitivity and Linearity |
|
Spike-in targets are mixes of ten synthetic transcripts that are spiked into the biological targets at known "Copies per Cell" equivalent with known "Red-to-Green ratios". The Multiple Linear Regression model is used to evaluate the overall system robustness and performance. A slope of 1.01 indicates a close-to perfect linear relationship between the two color channels1. The ability to accurately measure low copy number transcripts (as low as 0.5 to 1.5 copies per cell) reflects the unparallel sensitivity of the 60-mer oligo microarray. The correlation coefficient R2=0.9949 illustrates an overall robust performance of the platform. |
|
| Format-Independent Preservation of Sensitivity and Performance |
|
Differential expression experiments were performed on 22K Mouse Oligo microarrays (G4121A) and the data correlation was determined when compared to the 44K Whole Mouse Genome Oligo microarrays (G4122A). Log ratios derived from probes common to both microarrays are plotted against one another. The extremely low level of anti-correlation observed demonstrates the performance compatibility across the two formats. | 1 Data obtained using Feature Extraction Software (v.7.5) with default parameter settings: Spacial Detrending - ON,
Background Subtraction - OFF |