HiTS-FLIP
High-Throughput Sequencing With Fluorescent Ligand Interaction Profiling
HiTS-FLIP is a technique for measuring quantitative protein-DNA binding affinity at unprecedented depth. In this approach, the optics built into a high-throughput sequencer are used to visualize in vitro binding of a protein to sequenced DNA in a flow cell (Nutiu et al., 2011).
A microfluidic flow cell with anchored single-stranded DNA is sequenced by synthesis. Second-strand DNA is stripped and rebuilt using Klenow polymerase and unmodified dNTPs to form double-stranded DNA clusters. Fluorescently labeled binding protein is introduced at different concentrations, and binding is imaged.
Advantages:
- Quantitative and comprehensive
Disadvantages:
- Requires specialized hardware
- Not yet adopted widely by the scientific community
Reagents:
Illumina Library prep and Array Kit Selector
Reviews:
None available yet
References:
Nutiu R., Friedman R. C., Luo S., et al. Direct measurement of DNA affinity landscapes on a high-throughput sequencing instrument. Nat Biotechnol. 2011;29:659-664