What’s Inside The GeneAbacus?
GeneAbacus it is a ready-to-use reagent kit for in-house analysis of your gene editing experiment. It contains all buffers, enzymes and probes required to run the bioassay as well as the microfluidic chip for the simple digital quantification via fluorescence microscopy. Learn more below of how the COUNTed assay works.
Gene Editing Experiment
The gene editing experiment is performed by the customer. Countagen’s technology delivers a customized ready-to-use reagent kit for the analysis of such gene editing experiments.
Cell Lysis & DNA Fragmentation
DNA is extracted from the sample, either cells, tissue or organisms, such as Drosophila.
Specific Probing & Ligation
Tailored padlock probes are used to target the sequences of interest. Only in the event of a perfect match, a DNA ligase seals the nick and locks the padlock probe on the DNA target strand.
Amplification Via RCA
The circularized strands containing the target region are then amplified via Rolling Circle Amplification (RCA). In RCA, the circular template is replicated as a long single-stranded DNA concatemer that spools off when a strand-displacing DNA polymerase amplifies it. The resulting concatemeric amplicon consists of hundreds to thousands of repeats of the circular template.
The RCA amplicons, also termed RCA products, are labelled with sequence-specific fluorescent probes. The single-stranded RCA product collapses into a DNA micro-ball and due to the high negative charge stemming from the phosphate backbone it has minimal inter-molecular interactions. This single-molecule integrity makes RCA ideal for digital nucleic acid quantification.
RCA products can easily be detected using a standard fluorescence microscope and low magnification (10-20X). Countagen’s proprietary technology enriches all RCA products from solution into one field of view, which makes the quantification and analysis extremely simple. As shown in the scheme, RCA products are detected as bright fluorescent dots, where each dot represents a single DNA target from the beginning. This is the strength of RCA, being inherently digital without the need for compartmentalization as in digital PCR.
RCA provides the benefit of being inherently digital as one generated RCA product correspond to a single recognition event. This makes RCA a very powerful technology for digital quantification of nucleic acids with simple image analysis techniques.
As shown in the image on the left hand side, our technology offers in-house analysis of gene editing experiments with single nucleotide precision.
No specialized instrumention.
Ease of Use
Customized plug-and-play solution.
Highly accurate bioassay chemistry.
Assay stays in the hands of researchers.
Easily adaptable to other research questions.
A passionate team, ready to assist you.