STERIC FACTORS INFLUENCING HYBRIDIZATION OF NUCLEIC-ACIDS TO OLIGONUCLEOTIDE ARRAYS

We have investigated the use of spacer molecules to reduce steric inte rference of the support on the hybridisation behaviour of immobilised oligonucleotides, These spacers are built up from a variety of monomer ic units, using phosphoramidite chemistry, by condensation onto an ami ne-functionalised polypropylene support, The optimal spacer length was determined to be at least 40 atoms in length, giving up to 150-fold i ncrease in the yield of hybridisation, The effects of different charge d groups in the spacer were also examined, and it was shown that both positively and negatively charged groups in the spacer diminish the yi eld of hybridisation, Steric hindrance in hybridisation can also be a problem if the oligonucleotides attached to the support are too close to each other, Surface coverage was varied using a combination of clea vable and stable linkers, giving the highest hybridisation yields for surfaces containing similar to 50% of the maximum concentration of oli gonucleotides.