How Sequencing Works

LibraryA DNA or RNA sample that has adapters attached for sequencing. Preparation methods vary. is automatically denatured into single strands and further diluted onboard the instrument. During cluster generation, single DNA molecules are bound to the surface of the flow cell and amplified to form clustersA clonal group of DNA strands on a flow cell that produce one sequencing read. Each DNA strand on a flow cell seeds a template that is amplified until the cluster consists of hundreds or thousands of copies. For example, a flow cell with 10,000 clusters produces 10,000 single reads or 20,000 paired-end reads.. Cluster generation takes ~4 hours.

Clusters are imaged using two-channel chemistry (one green channel and one blue channel) to encode data for the four nucleotides. After one tile on the flow cell is imaged, the next tile is imaged. The process is repeated for each cycle of sequencing. Following image analysis, Real-Time Analysis software performs base callingDetermining a base (A, C, G, or T) for every cluster in a tile at a specific cycle., filtering, and quality scoring.Calculates a set of quality predictors for each base call, and then uses the predictor value to look up the Q-score.

As the run progresses, the control software automatically transfers concatenated base call filesContains the base call and associated quality score for every cluster of each sequencing cycle. (*.cbcl) to the specified output folder for data analysis. During the sequencing run, Real-Time Analysis software performs image analysis, and base calling. When sequencing is complete, secondary analysis begins. The method of secondary data analysis depends on your application and system configuration.

After initial primary analysis is complete, DRAGEN performs secondary analysis by using one of the available analysis pipelines in the selected analysis mode.