HOW MUCH WOULD THE PRESENCE OF AN ADDITIONAL SNP AFFECT DETECTION OF THE SNP OF INTEREST? HOW MANY ADDITIONAL SNPS COULD BE TOLERATED TO STILL ENABLE DETECTION OF THE SNP OF INTEREST BY HYB & SEQ?
If additional SNPs correspond to the middle of probe sequence, an increase in probe length could reduce the SNP effect on annealing. The probe length that we have successfully used ranges from 17 to 30 nt (Tm= 50 to 55oC) with a 10-T linker. The probe length and Tm could be increased to account for the internal SNP. Once annealing is achieved, the additional mismatches would not affect the discrimination of the terminal nucleotide, because it is based on ligation reaction rather than the strength of target hybridization.
Related Questions
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