Genpathway’s FactorPath™ identifies and quantifies the binding of transcription factors and co-regulating proteins such as co-activators and co-repressors to the genomic DNA within cells. Incorporating a technique called chromatin immunoprecipitation (ChIP), complexes comprising any protein involved in transcription and its associated DNA are isolated from cells and analyzed. Genpathway has developed optimized procedures for ChIP that yield maximum signal and minimal background, and also include important controls. Purification of DNA from the complexes results in the active DNA binding sites for the factor of interest. The DNA is analyzed using one or a combination of Genpathway’s analysis modes, including QueryMode™, which tests specific genomic regions for binding by quantitative PCR (Q-PCR); or DiscoveryMode™, Expanded Discovery™, ChIP-on Chip/ Tiling™, or ChIP-on Chip/Promoter, analysis modes that discover binding sites across the genome.

FactorPath—The Direct Method to Discover and Measure Gene Regulation

FactorPath measures the key events in the regulation of gene transcription—the primary level at which compounds and other changes exert their effects within cells. FactorPath can detect changes in protein binding within a few minutes—a capability that RNA profiling cannot provide. FactorPath determines how genes are activated or repressed by transcription factors and coregulating proteins and can help identify the primary targets of compounds versus secondary and later targets. FactorPath determines absolute factor-binding levels, including baseline levels, which can be compared across genes, cells, treatments, experiments, and time. Because Genpathway’s assays measure DNA, which is more stable than RNA, the transcription complexes analyzed are more durable in cells and tissues.

Advantages of FactorPath in Analyzing Gene Regulation

Genpathway’s direct gene regulation assays are accurate, sensitive, quantitative, and highly reproducible. Repeat experiments involving cells and animals treated months apart show essentially identical changes in factor binding. Transcription complexes are very stable, such that tissue held under suboptimal conditions—for example, at room temperature for hours or at -70°C for years—can be assayed. Tissues that contain high levels of RNase, such as pancreas, and unseparated whole blood can be assayed.

FactorPath Analyzes a Different Endpoint than Gene Expression Profiling

FactorPath identifies the genomic regions functionally bound (inside living cells) by specific transcription factors and co-regulating factors, and quantifies their binding levels. This information can be used to define and even discover the genes that are being regulated and undergoing differential expression. Knowledge about transcription factor-binding activities also provides information about how genes are regulated and co-regulated in response to changes such as treatment with a drug. The ability to look at functional sites with ChIP, as opposed to all sites containing consensus sequences, is critical because only a fraction of potential binding sites are used in cells.

Applications of FactorPath

The types of applications are extensive:

Genpathway Offers Complete ChIP Assay Services

Genpathway’s transcription-based assays include FactorPath™, which directly analyzes how genes are regulated by transcription factors and co-regulating factors; and TranscriptionPath™, which directly analyzes the process of transcription. In many cases, a combination of these two types of assays that correlates changes in gene regulation and transcription will provide the most valuable information. Genpathway offers both types of assays as a service with any of four analysis modes, with a focus on high-quality data, rapid turnaround, and cost effectiveness. Customers supply the samples to be tested—cells that have been briefly fixed, or frozen tissue—along with assay-specific information.