Advantages of the SNP Array

The SNP array has several advantages over traditional methods such as karyotyping and allelotyping with microsatellite markers. First, it requires no cell cultures, eliminating some of the karyotyping problems that arise from passaging cells. Cultures from primary tumors are often contaminated with normal cells, which can proliferate and thus complicate the karyotyping results. Second, the SNP array has a high SNP density for genotyping and copy number measurements. For example, on the GeneChip human mapping 500 K array, the median physical distance between SNPs is 2.5 kb, and the mean distance is 5.8 kb. Eighty-five percent of the human genome is within 10 kb of a SNP, which provides a mapping resolution that is 100- to 300-fold higher than the current panels of ~400 short tandem repeat markers (18). Moreover, the mean heterozygosity of these SNPs is 0.30, which generates many heterozygous informative genotype calls for inferring the LOH region.

The SNP array has been shown to be highly accurate (99.5%) and reproducible (99.9%) and has a high call rate (95.0%) in genotyping (23). It can also be used to detect changes in the chromosome copy number at a single SNP locus resolution, as each SNP locus is analyzed with 40 SNP probes. The hybridization signals of these SNP probes depend on the copy numbers of DNA at each SNP locus. A statistical model has been developed that calculates the likelihood of determining LOH, SNP copy number changes, and chromosome copy number changes in one analysis without using normal DNA as the reference (43,44). This model can reliably detect amplifications and homozy-gous deletions that extend over regions of less than 1 Mb. In several cancer studies, it has been used to successfully determine LOH in newly identified and previously known chromosome regions (2,43,45,46,47,48,49). Figure 1 illustrates a simultaneous detection of copy number changes and LOH using the 10 K SNP array, as presented in our recent paper (49).

The ability of the SNP array analysis to detect copy number changes and LOH simultaneously from a tumor sample has an added advantage over CGH. In several studies, different LOH mechanisms (i.e., LOHs with and without copy number changes) have been identified using SNP arrays. LOH detection with an SNP array analysis is based on genotyping calls and can thus detect the loss of one allele, followed by the reduplication of the remaining allele as LOH. We have found that amplified EGFR and PDGFRa genes are located in regions of LOH (Fig. 2), which implies that a specific allele is amplified (49). A sequencing analysis of the EGFR transcript in the tumor confirmed that the amplified EGFR allele carried a type III mutation and was the only amplified allele in the LOH region. This may represent a new mechanism of tumor progression (49). Similarly, another recent SNP array study of 100 cases of lung cancer revealed

Fig. 1. Side-by-side whole-genome LOH patterns and copy number changes in 28 pediatric gliomas. Top: The sample clustering tree is based on LOH data in the significant regions. LOH regions in each tumor are highlighted in dark gray color. Bottom: A darker color indicates a higher copy numbers at the corresponding regions. The graph was generated by dChip2006 software. (Reprinted with permission of the American Association for Cancer Research from Wong et al., Cancer Res 2006; 66(23): 11172-11178).

Fig. 1. Side-by-side whole-genome LOH patterns and copy number changes in 28 pediatric gliomas. Top: The sample clustering tree is based on LOH data in the significant regions. LOH regions in each tumor are highlighted in dark gray color. Bottom: A darker color indicates a higher copy numbers at the corresponding regions. The graph was generated by dChip2006 software. (Reprinted with permission of the American Association for Cancer Research from Wong et al., Cancer Res 2006; 66(23): 11172-11178).

Allelic imbalance in pediatric gliomas

Allelic imbalance in pediatric gliomas

Fig. 2. An enlarged region of Fig. 1 showing amplification of specific alleles in LOH regions (49).

that DNA amplification is derived from a single allele or the remaining allele located at LOH region (50). In contrast, CGH is based on the ratio of the DNA copy number to a reference DNA and thus cannot detect reduplication of the remaining allele in the case of LOH, which will be detected as no change.

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