Human SNP Kits

    Ecoli s.r.o. offers wide range of SNP kits, based on pyrosequencing technology, for rapid and cost-effective analyzis of human DNA in clinical samples. Pyrosequencing technology is a unique method for short-read DNA sequencing and mutation or SNP analysis. It is suitable for applied genomics including molecular applications for disease diagnosis, clinical prognosis and pharmacogenomics testing.

 

Principle

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  Pyrosequencing is based on the detection of released pyrophosphate (PPi) during DNA synthesis. In a cascade of enzymatic reactions, visible light is generated that is proportional to the number of incorpo-rated nucleotides. The cascade starts with a nucleic acid polymerization reaction in which inorganic PPi is released as a result of nucleotide incorporation by polymerase. The released PPi is sub-sequently converted to ATP by ATP sulfurylase, which provides the energy to luciferase to oxidize luciferin and generate light. Because the added nucleotide is known, the sequence of the template can be determined. When the light signal is detected, the base is registered and the next nucleotide is added. If the added nucleotide is not complementary to the next base in the template, no light is generated.

 

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     There are two different pyrosequencing strategies that are currently available: solid-phase pyrosequencing and liquid-phase    pyrosequencing. Solid-phase pyrosequencing utilizes immobilized DNA in the three-enzyme system described previously. In this system a washing step is performed to remove the excess substrate after each nucleotide addition. In liquid-phase pyrosequencing apyrase, a nucleotide-degrading enzyme from potato, is introduced to make a four-enzyme system. Addition of this enzyme eliminates the need for solid support and intermediate washing thereby enabling the pyrosequencing reaction to be performed in a single tube.

 

     The combination of instrumentation, dedicated software and reagent kits make pyrosequencing technology ideal for analysis of all genetic diversities such as bi-tri- and tetra-allelic polymorphisms, multiple SNPs, mutations and insertions/deletions (InDels).

     Pyrosequencing is unique among genotyping methods in that the measurement of every allele is fully quantitative. This property has made pyrosequencing a primary choice for SNP screening in DNA pools, quantification of the degree of DNA /CpG methylation in epigenetic research, the analysis of hematopoeitic chimerism and discriminating between mixed genotypes in heterogeneous samples (e.g. tumor and normal cells). Because both alleles are extracted and measured in a single sample, this method is insensitive to differences in extraction efficiency and eliminates the need for control genes or quantification of total RNA recovery. Samples for pyrosequencing detection can be blood, tissue or cells collected on a swab.

 

Typical manuals for SNP kits:

    BRCA-screen-LA

    FOLATE-screen-LA

Real TimeFEP KitsElectrophoresis KitsAla 1/4 Detector