Hundreds of copies of the genome are stretched simultaneously on one substrate
Single step stretching and assembly of millions of large DNA fragments
Molecular Combing is a user-friendly technique for the direct visualization of DNA at the single molecule level. The Molecular Combing System exploits surface forces to simultaneously stretch and assemble millions of DNA molecules on the surface of a vinyl silane treated glass carrier. The process is simple, fast and applicable to DNA molecular sizes exceeding the megabase. The technique is compatible with pre-labeled DNA and with in situ hybridization. No amplification is needed in the process; therefore no bias is introduced in the DNA population to be analyzed. Moreover, a large number of genome copies can be combed per experiment, this provides a statistically relevant number of measurements on the stretched DNA.
To implement the molecular combing technique, Genomic Vision developed the Molecular Combing System, an automatized instrument that stretches DNA molecules onto specially-treated glass surfaces (CombiCoverslips) in a 10-minute hand-off procedure. The resulting DNA is organized in discrete strands and can be analyzed at single molecule level.
Constant stretching factor along each molecule (1 μm ≈ 2 kb)
Allows for accurate and reproducible measurements for physical mapping of loci of interest
- Uniform stretching for all molecules over the surface: measurements can be pooled without local calibration
- Stretching insensitive to sequence content: no GC content bias, all genomic regions get stretched and are equally represented on the slide enabling the study of any regions of interest
- Unbiased stretching irrespective of the size of the DNA molecules: the longer the extracted molecule, the wider the view for the analysis of the region of interest
- Hundreds of copies of the genome are stretched simultaneously on one substrate: offers a unique sensitivity to rare events, heterogeneous samples, mosaicism and enables precise statistical analysis.
The Molecular Combing procedure
High molecular weight DNA is extracted and purified from the samples, and the DNA solution is placed in the disposable reservoir of the MCS. A 22-mm2 CombiCoverslip (glass carrier specifically treated with vinyl silane) is manually clipped to the system. The rest of the procedure is fully automated and takes only 10 minutes.
First, the CombiCoverslip is lowered into the reservoir containing the DNA, in which it is incubated briefly to allow the DNA to attach to the glass surface at one or both of its extremities. The CombiCoverslip is then automatically withdrawn at constant speed from the DNA solution. Due to the slide being drawn through the air-liquid meniscus, DNA molecules are stretched and aligned in a controlled and consistent manner on the glass surface. Each DNA molecule is combed out with a constant stretching factor (2 kb/µm), enabling precise and direct measurements along the length of the DNA fiber.