Survey Entire Microbial Genomes with Unmatched Speed

With the current capacity of 385,000 custom probes, the initial survey of a microbial genome can span up to 1,300,000bp of genomic sequence in a single array. For larger genomes, the design can be split over as many arrays as required for complete genomic coverage. In the first phase of CGS, genomic alterations are identified by labeling test and reference genomic DNA samples and co-hybridizing to survey arrays derived from both strands of the reference genome. The locations of genomic alterations are identified in a single round of hybridization (see Figure 1).

Rapid, Targeted Resequencing of Regions of Interest

The genomic regions of interest identified in the first phase provide the content for targeted sequencing arrays in the second phase of CGS. Targeted arrays, sequencing only the regions of the genome where alterations exist, maximize the yield of useful data from every synthesized array. Because NimbleGen's highly flexible Maskless Array Synthesis (MAS) technology enables rapid synthesis of new array designs, the results of phase one are quickly incorporated into sequencing array designs in phase two for the characterization of microbial mutations.

Accurate, Scalable SNP Detection

NimbleGen’s CGS technology is the most efficient method for the rapid localization and characterization of genomic alterations on a genome-wide scale. Efficiently detecting genomic changes, CGS can sensitively characterize even a single base change in an entire genome. In one study, CGS utilized only five arrays to effectively identify the only single base change in a 5,000,000bp genome, generating zero false positives. CGS typically identifies ~95% of SNPs present in unique regions of genomes, with less than one false positive per 100,000 to 1,000,000 bases analyzed. CGS also readily identifies regions requiring manual sequencing, such as insertions and deletions, thus greatly simplifying the process of fully characterizing most genomic alterations.

CGS has a wide range of applications in microbial comparative genomics, including:

• Characterizing industrially important microbes for food, pharmaceutical, and chemical industries.
• Characterizing genomic alterations associated with virulence and attenuation.
• Genotyping infectious pathogens for strain identification.
• Tracking genomic alterations associated with environmental changes in natural environments and chemostat/ fed batch settings.
• Rapidly characterizing phenotypically interesting microbes.
• Identifying genes responsible for anti-microbial agent resistance.

Phase 1. Mutation Mapping

1. Sample Fragmentation: The test and reference DNA samples are separately cleaved to pools of low molecular weight fragments.
2. Sample Labeling: Test and reference samples are independently labeled with fluorescent dyes, Cy3 and Cy5 respectively.
3. Hybridization: Test and reference samples are hybridized to a NimbleGen CGS whole-genome tiling array.
4. Data Analysis: Data is extracted using NimbleScan software, the ratio of reference versus test sample are plotted versus their genomic position.

Phase 2. Resequencing

5. Sequencing Array: The genomic sequence surrounding each identified ratio peak is converted into a sequencing array querying base positions surrounding each peak on both DNA strands.
6. Hybridization: The labeled test DNA is hybridized to the sequencing array.
7. Data Analysis: Data is extracted using NimbleScan software. The resulting sequences are compared to the reference sequence and SNPs are called and categorized as synonymous, non-synonymous, or non-coding. Regions of ambiguous calls are flagged by genomic position for further characterization.

Brochures & Datasheets

• NimbleGen CGS Microarrays and Services
  Brochure (PDF Format 1.2MB)

For a complete listing of literature covering all Roche NimbleGen products and services please visit our literature page.