Genetics

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“MOLgen Genetics Interleukin 28B Kit” is an Assay Kit for the determination of single nucleotide polymorphisms of interleukin 28B gene using real-time polymerase chain reaction with detection of melting curves.

“MOLgen Genetics Interleukin 28B Kit” is designed for the determination of single nucleotide polymorphisms of interleukin 28B gene using real-time polymerase chain reaction with detection of melting curves. The kitis intended for the differential determination of single nucleotide polymorphisms rs12979860 C/T and rs8099917 T/G of interleukin 28B gene (IL28B) using polymerase chain reaction (PCR) with melting of hybridisation products with a fluorescent tag and analysis of melting curves.

The extraction of DNA from clinical material can be performed using the kit: “MOLgen Universal Extraction Kit”.

The kit is intended for use with real-time PCR cycler CFX96 (Bio-Rad, USA).

The kit is designed to test 48 samples, including control samples.

The principle of the method is based on the amplification of the selected human DNA region followed by detection of melting curves of hybrid complexes in PCR products and specific probes. The process of amplification consists of repeated cycles: temperature denaturation of the DNA matrix, annealing primers with complementary sequences of the DNA matrix, and synthesis of a complementary chain from these primers with Taq polymerase.

After the amplification reaction, the lowering of temperature results in hybridisation of the specific DNA probe with the amplification product and a decrease in the fluorescent signal in the tube. During melting of formed duplexes, a DNA probe bearing a fluorescent dye is released. With incomplete complementarity of the DNA probe and the selected DNA region, the melting temperature of the duplex is lower than with complete complementarity. Thus, the melting temperature of probe duplexes and amplification products is different for two allelic variants. As a result, the genotype for each of the polymorphisms to be determined can be identified.

Determination of single nucleotide polymorphisms rs12979860 C/T and rs8099917 T/G of IL28B gene in patients infected with genotype 1 hepatitis C virus has prognostic value when choosing a treatment plan and predicting the disease outcome.

The genotype rs12979860 С/С increases the probability of positive response when treating a patient with Interferon and Ribavirin, and also increases the probability of spontaneous viral clearance.

The genotype rs8099917 Т/Т increases the probability of spontaneous viral clearance irrespective of treatment. The genotype rs8099917 G/G increases the risk of low response when treating a patient with pegylated Interferon and Ribavirin.