The therascreen KRAS RGQ PCR Kit is intended to detect 7 mutations in codons 12 and 13 of the KRAS gene. The KRAS gene encodes a protein that plays a critical role in the epidermal growth factor receptor (EGFR) signaling cascade. Mutations in the KRAS gene can affect how the protein stimulates these downstream pathways. KRAS is mutated in approximately 30% of all cancer types. Cancers that exhibit a high frequency of KRAS mutation include colorectal cancer (35%) and lung (18%) cancer.
KRAS mutations detected by the therascreen KRAS RGQ PCR Kit include:
- 13 ASP
The kit utilizes 2 technologies — ARMS (Amplification Refractory Mutation System) and Scorpions — for detection of mutations in real-time PCR.
Allele- or mutation-specific amplification is achieved by ARMS. Taq DNA polymerase is effective at distinguishing between a match and a mismatch at the 3' end of a PCR primer. Specific mutated sequences are selectively amplified, even in samples where the majority of the sequences do not carry the mutation. When the primer is fully matched, the amplification proceeds with full efficiency. When the 3' base is mismatched, only low-level background amplification occurs.
Detection of amplification is performed using Scorpions. Scorpions are bifunctional molecules containing a PCR primer covalently linked to a probe. The fluorophore in this probe interacts with a quencher, also incorporated into the probe that reduces fluorescence. When the probe binds to the amplicon during PCR, the fluorophore and quencher become separated. This leads to an increase in fluorescence from the reaction tube.