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Studies of Surface Hybridization In a Model System

Studies of Surface Hybridization In a Model System Ping Gong, Damion Irving, Dongbiao Ge , Rastislav Levicky Polytechnic Institute of New York University.

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Studies of Surface Hybridization In a Model System

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  1. Studies of Surface Hybridization In a Model System Ping Gong, Damion Irving, DongbiaoGe, RastislavLevicky Polytechnic Institute of New York University This research was supported by the National Institutes of Health (HG003089 & HG004512) and by National Science Foundation (DMR 07-06170).

  2. Noncompetitive Hybridization Surface Hybridization Regimes • easy • highly reproducible • preserves probe activity HO-(CH2)3-SH DNA-SH a “DNA brush” Gold support thiol or disulfide * Herne T.M. & Tarlov M.J. JACS 1997 20mer Probe: 5’-TTTTAAATTCTGCAAGTGAT – (CH2)3-S-S-(CH2)3-OH -3’ 18mer Target: 3’-AATTTAAGACGTTCACTA-5’ 7 probe Coverages (S0) 5 ionic strengths (CB) CT = 100 nM

  3. Surface Hybridization Regimes L: Langmuir x = SD/S0 S0 (1012 cm-2) Gong, Levicky PNAS 2008 Irving, Gong, Levicky JPCB 2010 Gong, Wang, Liu, Shepard, Levicky JACS 2010

  4. Surface Hybridization Regimes PL: Pseudo-Langmuir x = SD/S0 S0 (1012 cm-2) surface DGSo = -44,000 kJ/mol vs -100,000 kJ/mol in solution (reference state 1 M Na+, 22 C) Gong, Levicky PNAS 2008 Irving, Gong, Levicky JPCB 2010 Gong, Wang, Liu, Shepard, Levicky JACS 2010

  5. Surface Hybridization Regimes SH-E:Electrostatically suppressed hybridization x = SD/S0 S0 (1012 cm-2) Gong, Levicky PNAS 2008 Irving, Gong, Levicky JPCB 2010 Gong, Wang, Liu, Shepard, Levicky JACS 2010

  6. Surface Hybridization Regimes SH-P: “Packing” suppressed hybridization x = SD/S0 S0 (1012 cm-2) CB = 0.63 mol L-1 Gong, Levicky PNAS 2008 Irving, Gong, Levicky JPCB 2010 Gong, Wang, Liu, Shepard, Levicky JACS 2010

  7. Competitive Hybridization 20mer Probe: 5' HS-(CH2)6-CAG GCA CAA ACA TGA ACC TC 3‘ 20mer Perfect Match (PM) Target: 5' GAG GTT CAT GTT TGT GCC TG-(CH2)3-NH2 3‘ 20mer Mismatch (MM) Target: 5' GAG GTT CGT GTT TGT GCC TG3' 0.2 M PB pH 7.0 10:1 MM:PM range of temperatures Thermally-stable Surface Immobilization DNA

  8. Solution Melt Transitions (o) Perfect match: DHPMo, DSPMo Mismatch: DHMMo, DSMMo Mismatch Penalty in Solution: DDHo = DHMMo - DHPMo DDSo = DSMMo - DSPMo PM Surface Melt Transition (*) Perfect match: DHPM*, DSPM* assume mismatch penalty same as in solution: DHMM*= DHPM* + DDHo DSMM*= DSPM* + DDSo

  9. 20mer Probe: 5' HS-(CH2)6-CAG GCA CAA ACA TGA ACC TC 3‘ 20mer Perfect Match (PM) Target: 5' GAG GTT CAT GTT TGT GCC TG-(CH2)3-NH2 3‘ 20mer Mismatch (MM) Target: 5' GAG GTT CGT GTT TGT GCC TG3' 0.2 M PB pH 7.0 10:1 MM:PM ● max discrimination ~ 20 C below solution TM for PM ● suppression of PM signal at lower temperatures ● enhancement of PM signal at higher temperatures

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