Increasing detection of polymerase chain reaction (PCR) by isolation of PCR products (IPCRp).

AIM

To develop a method for enhanced polymerase chain reaction (PCR) product detection.

METHODS

During the PCR, the double-stranded product is generated with fluorescent dye on one strand, and biotin on the other strand. The product is captured on the streptavidin-coated plates with high efficiency (IPCRp). Washing of the all unamplified compounds, including dye-labeled unincorporated primers, follows the PCR. The targeted dye-labeled PCR product is released by denaturation and loaded on the detection platform.

RESULTS

After the application of the IPCRp, the resulting product is highly concentrated targeted dye-labeled single-stranded DNA, free of the unincorporated primers and other PCR artifacts. The strength of the signal of the IPCRp product on detection platform is two- to five-fold higher than the strength of the signal of the conventional PCR product.

CONCLUSION

The IPCRp procedure can be accomplished in less than 20 minutes. Efficient isolation of the PCR products has two steps, washing and denaturation. It can increase the yield of targeted PCR product and increase the sensitivity of the detection platform.