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DirectBlood Genotyping PCR Kit

  • DirectBlood Genotyping PCR Kit is a novelty on the market. It allows genotyping in real-time and without any previous DNA isolation. You simply save and money: directly use EDTA blood without any extraction steps.
    The DirectBlood Genotyping PCR Kit is optimized to function with a wide range of different SNPs to detect possible nucleotide variations. It has been successfully tested with a variety of hydrolysis probes and hybridization probes.

    The setup of a genotyping experiment can not be simpler anymore: Mix primers&probes with the Genotying mastermix, add the respective blood sample and start real-time PCR, this is it. 

    It is even more simple: store the DirectBlood Genotyping PCR Kit at room-temperature. A cool chain is not needed anymore. The included master mix is freeze-dried and dissolves within a few seconds after addition of the rehydration buffer.

    Additionally, we have already established SNP-assays for you. In case, you can not find your respective SNPs, please provide us with the rs-number of the SNP-assays your are interested-in and you will receive an individual quotation from us.

  • DirectBlood Genotyping PCR Kit contains all components necessary for direct PCR or PCR based genotyping directly from blood samples including an engineered DNA polymerase, an specifically optimized reaction buffer system, and ultrapure dNTPs. An aptamer-based hot-start formulation of the included blood-resistant DNA polymerase prevents false amplification. Temperatures above 50°-55°C cause the aptamer’s secondary structure to melt and will set-free the polymerase. Only target specific primers and probes need to be added.

  • The freeze-dried DirectBlood Genotyping PCR Lyocake can be stored at room-temperature. Please store resuspended master mixes (2x) and primer/probe mixes (4x) at -20°C. Minimize the number of freeze-thaw cycles by storing in aliquots. For a day-to-day use, we recommend keeping an aliquot at 4°C. Please store the included rehydration buffer and additional primer/probe mixes upon arrival at -20°C.

DirectBlood Genotyping Kit (#5000) performance with a hydrolysis probe assay

The DirectBlood Genotyping Kit is compatible with a wide range of assays: Here we are detecting a SNP in the Factor V gene (rs6025; G1691A), which is connected with a higher risk for venous thrombosis, with a fluorescent hydrolysis probe assay.

Homozygous wild type blood samples will show a strong signal and amplification plot (blue curve) in channel 2 (Cy5) and none or a very low signal

(red curve) in the other channel 1 (ROX).

 

 

Heterozygous blood samples will give an intermediate signal and amplification plots with very similar intensities in both channels 1 and 2 (blue and red curve).    Homozygous mutant blood samples, channel 1 (ROX) will show a strong signal and amplification plot (red curve) and channel 2 (Cy5) none or a very low signal and amplification plot (blue curve). So the signals are reversed on homozygous mutant samples.   


DirectBlood Genotyping Kit performance with a hybridization probe assay

Here we are detecting the same SNP in the Factor V gene (rs6025; G1691A), which is connected with a higher risk for venous thrombosis, with a fluorescent hybridization probe assay.

Three blood samples with known genotype were used. All the genotypes can clearly be identified. The blue curve shows the specific wild type (homozygous G;G) peak at 61°C. The red curve shows the specific mutant (homozygous A;A) peak at 53°C. A heterozygous case (A;G) will result in both melting curve peaks, respectively at the same specific temperature for wild type and mutant (orange curve).


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