iGenomX library construction products are purpose built for each application. The ideal data set for an application is envisioned, then molecular methods are developed to achieve the ideal.
Each iGenomX application builds from a straightforward, low cost, fully automatable methodology centered around polymerase. These polymerase enzymes catalyze the formation of long-chain molecules by linking smaller molecular units, such as nucleotides with nucleic acids. Uncontrolled polymerization causes amplification bias and error propagation, which leads to increased costs and reduced data quality. Proprietary iGenomX library construction empowers researchers with precise control of polymerase activity. It starts with a simple workflow:
- P5 Adapter tailed random primers hybridize to template DNA. Polymerase extends and terminates polymerization by incorporation of a biotinylated ddNTP. The ratio of ddNTP / dNTP controls the length of initial priming events.
- Terminal biotin polymers are captured on streptavidin coated magnetic beads and washed to remove excess primers, NTPs and polymerase. P7 adapter tailed random primers hybridize to captured molecules and are extended with the use of a strand displacing polymerase. The primer closest to bead remains bound and displaced product is washed away.
- PCR amplification with full length sequencing adapters occurs directly on beads. Library products are size selected and ready for sequencing.
Cost savings result from:
- No chemical or physical fragmentation
- No blunt end repair
- No A-tail
- No ligation
- Polymerase is only enzyme used