Nucleic Acids Research, Vol 25, Issue 6 1155-1161, Copyright © 1997 by Oxford University Press
MS Shchepinov, SC Case-Green and EM Southern
We have investigated the use of spacer molecules to reduce steric
interference of the support on the hybridisation behaviour of immobilised
oligonucleotides. These spacers are built up from a variety of monomeric
units, using phosphoramidite chemistry, by condensation onto an
amine-functionalised polypropylene support. The optimal spacer length was
determined to be at least 40 atoms in length, giving up to 150-fold
increase in the yield of hybridisation. The effects of different charged
groups in the spacer were also examined, and it was shown that both
positively and negatively charged groups in the spacer diminish the yield
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 cleavable and
stable linkers, giving the highest hybridisation yields for surfaces
containing approximately 50% of the maximum concentration of
oligonucleotides.
ARTICLES
Steric factors influencing hybridisation of nucleic acids to oligonucleotide arrays
Department of Biochemistry, University of Oxford, South Parks Road Oxford OX1 3QU, UK. misha@bioch.ox.ac.uk
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