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DNA origami attachment and AFM imaging on mica and SiO 2 /Si [100] in air

DNA origami attachment and AFM imaging on mica and SiO 2 /Si [100] in air. Foundations of Nanoscience (FNANO2010) NSF workshop on DNA origami Kyoung Nan Kim kkim4@nd.edu University of Notre Dame 042610. Outline. Self-assembly of DNA origami

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DNA origami attachment and AFM imaging on mica and SiO 2 /Si [100] in air

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  1. DNA origami attachment and AFM imaging on mica and SiO2/Si [100] in air Foundations of Nanoscience (FNANO2010) NSF workshop on DNA origami Kyoung Nan Kim kkim4@nd.edu University of Notre Dame 042610

  2. Outline • Self-assembly of DNA origami • DNA origami attachment and Tapping mode AFM imaging on mica in air • DNA origami attachment and Tapping mode AFM imaging on SiO2/Si [100] in air - Preparing clean and smooth SiO2/Si [100] - Cationic self-assembled monolayers (SAMs) on SiO2/Si [100]

  3. Self-assembly of rectangular DNA origami 90 °C for 5 min in thermal cycler b)20 °C ( -1 °C/min) c) Kept at 4 °C M13mp18 + 20 - 100 fold excess helper strands in TAE/Mg2+(1x, pH8) Bayou Biolabs Integrated DNA technologies 2 nm hight Purification Microcon YM-100 Centrifugal Filter Unit,100K MWCO, Cat.# : 42413 Millipore 70 nm DNA origami Soln. 90 nm Rothemund, P. W. K.; Nature 2006, 440, 297-302

  4. DNA origami attachment and Tapping mode AFM imaging on mica in air

  5. What is mica? • Thin, flexible, and transparent • Micaceous cleavage / Easily cleaves layer by layer • The surface is negatively charged http://chemistry.binghamton.edu/ZHONG/spm/stmafm1.htm (accessed on 04/09/10)

  6. Introduction / DNA attachment on mica DNA Mg2+ Mg2+ TAE/Mg2+ (Sterile, 1x, pH 8) • Tris : 40 mM • Acetic Acid : 20mM • EDTA : 2mM • MgCl2 : 12.5 mM O O O O

  7. DNA origami attachment on mica • Mica (Electron Microscopy Sciences, Muscovite Mica V-5 (2" x 3"), Thickness :0.15 -0.21mm, Cat. # : 71850-01 (10/pk)): Top layer is carefully removed by scotch tape and DNA origami solution is directly applied on freshly cleaved mica • DNA origami deposition - 5 µl of 3 nM DNA origami (1x TAE/Mg2+, pH 8) is deposited on freshly cleaved mica - Deposition time : around 30 sec. up to 1 min. - The surface is rinsed with 18MΩ water and dried with N2 gas

  8. Various deposition conditions • 30 second, 100x helper Deposition time • 3 min 3 nM, 20 x helper 6 nM Concentration Of DNA origami

  9. AFM imaging of DNA origami on mica in air • Multimode NanoscopeIIIa from Veeco instruments Inc. • Tapping mode AFM imaging in air • AFM probes (non-contact/tapping mode in air)- NSG30-W from NT-MDT (410 chips in a wafer, Au reflective coating) and T300-W from Vista probes (410 chips in a wafer, bare) a. Resonant frequency (nominal): 300kHz b. Force constant: 40 N/m c. Tip radius : < 10 nm • Image analysis : WSxM 5.0 Develop 1.0 (free software) http://www.nanotec.es/products/wsxm/ I. Horcas et al. Rev. Sci. Instrum. 78, 013705 (2007)

  10. Tapping mode AFM image of DNA origami in air Scan size: 5 um2 Scan speed: ~ 1.0 Hz Samples/line: 512 Set points: 1.0-1.2 V Integral gain: 0.2 Proportional gain: 0.4 Scanning time: ~ 4 min.

  11. 60.065 nm 90.618 nm

  12. DNA origami attachment and Tapping mode AFM imaging on SiO2/Si [100] in air

  13. DNA attachment on cationic SAMs on SiO2/Si [100] DNA DNA Mg2+ Mg2+ Mg2+ Mg2+ 3-aminopropyltriethoxysilane (APTES) trimethylaminopropyltrimethoxysilyl chloride (TMAC) NH2 NH3+ NH3+ N(CH3)3+ N(CH3)3+ N(CH3)3+ Si Si Si Si Si O O Si O O O O O O O O O n O n Si Si Si Si Si Si SiO2 Si [100] SiO2 Si [100]

  14. Preparation of clean silicon surface • Piranha cleaning - Silicon chip (MEMC Electronic Materials, Inc., Malaysia) is soaked in piranha solution (H2O2:H2SO4=1:3) at 70 °C for 30 min. Caution: Piranha solution is a strong oxidant and can cause explosions when mixed with organic solvents! • RCA cleaning - HF treatment : HF (10 %) is stored in PTFE beaker. HF etches SiO2 and surface becomes smooth and hydrophobic - RCA 1 treatment (NH4OH:H2O2:H2O=1:1:50) : Removes organic residues at ~ 70 °C for 10 min. - RCA 2 treatment (HCl:H2O2:H2O = 1:1:50): Removes metallic impurities at ~ 70 °C for 10 min. - Stored in 18MΩwater for the long term storage

  15. RCA cleaning bench Basket RCA bath HF bath Nitrile gloves Apron, face shield Waste bottles C:\Users\Kyoung Nan Kim\AppData\Local\Temp\Temp1_MSDs_HF.zip\H3994.htm

  16. Tapping mode AFM image of silicon chip before the cleaning, and after Clean SiO2 after RCA cleaning Dirty SiO2 RMS: 0.1084 nm • RMS: 3.2786 nm

  17. Preparation of cationic SAMs on SiO2 1% APTES 1% APTES • 3-aminopropyltriethoxysilane (APTES, stored in N2, Gelest inc., Cat. #: SIA0610.0) and N-trimethoxysilylpropyl-N,N,N,-trimethyl-ammonium chloride (TMAC, 50% in MeOH, stored in N2, Gelest inc., Cat. #: SIT8415.0-25GM ) - APTES and TMAC solution is stored at 4 °C - APTES and TMAC polymerizes in water • APTES SAMs deposition - RCA cleaned silicon chip is soaked in 1-2% APTES and TMAC solution for 30 min. in 18MΩ water - After the deposition, the silicon chip is sonicated in MC for 10 min. to remove physisorbed APTES residues - APTES and TMAC treated silicon chip is stored under 18MΩ water for the long term storage Old bottle New bottle RMS: 0.7492 nm RMS: 0.1133 nm 1% TMAC 2% TMAC RMS: 0.2712 nm RMS: 0.1537nm

  18. T-AFM image of bare SiO2 and APTES/SiO2 Bare SiO2 1% APTES on SiO2 (30min soaking) RMS: 0.1133 nm Contact angle: 65.8 ° RMS: 0.1084 nm Contact angle: NA Characterization of cationic SAMs on SiO2 • X-ray photoelectron spectroscopy (XPS) • Tapping mode AFM in air • Contact angle measurement

  19. Tapping mode AFM image of 1% APTES in various soaking time 10 min 20 min 30 min RMS (Rq): 0.2405 nm 0.1296 nm 0.1133 nm Contact angle: Avr 62.67 ° ± 5° 55.8 ° 65.8 ° 40 min 50 min 60 min RMS (Rq): 0.3065 nm 0.3573 nm 0.1944 nm Contact angle: 60.25 ° 62.5 ° 62.67 °

  20. T-AFM image of DNA origami on 1% APTES/SiO2

  21. T-AFM image of DNA origami on 2% TMAC/SiO2 90.913 nm 40.709 nm

  22. Time dependent multi-scanning in airDNA origami on 2% TMAC (a) (b) (d) (c)

  23. Selective Binding of DNA on Silicon Koshala Sarveswaran University of Notre Dame

  24. Step 1. Cleaning the silicon chip SiO2 Silicon(100) Freshly cleaned silicon chip with a thin layer of oxide (native oxide)

  25. AFM Image Freshly cleaned silicon substrate with native oxide

  26. E-beam Resist Polymethylmethacrylate (PMMA) Polymethylmethacrylate (PMMA) was one of the first materials developed for e-beam lithography. It is the standard positive e-beam resist and remains one of the highest resolution resists available. PMMA is usually purchased in two high molecular weight forms (496 K or 950 K) in a casting solvent such as chlorobenzene or anisole.

  27. PMMA vendor: MICRO CHEM Corp. 90 Oak St. Newton, MA 02464 2% PMMA (950 K) in Anisole 500 ml ---$ 364 (http://www.microchem.com/products/pdf/PMMA_Data_Sheet.pdf)

  28. 2. Spinning the resist (PMMA) on silicon • Use a clean dry silicon sample • Select the recipe on the spinner • Speed 4000 rpm time 30 sec. • Bake on a hotplate 1800C for 2-3 minutes. • or bake in the oven at 1800C for 5 hrs. 80-100 nm thick PMMA on the silicon

  29. AFM Image PMMA on silicon

  30. e-beam e-beam 3. Electron beam lithography (EBL) Expose at 75 keV Dose 600-800 µC/cm2 Elionix ELS 7000 system PMMA Electron beam resists are the recording and transfer media for e-beam lithography. electron exposure modifies the resist, leaving it either more soluble (positive) or less soluble (negative) in developer. Silicon (100)

  31. PMMA after e-beam exposure PMMA Exposed PMMA Silicon(100) -PMMA is a positive resist -exposed regions contain very soluble fragments

  32. 4. Development after EBL Prepare developer methyl isobutyl ketone:isopropanol (MIBK:IPA 1:3) or methyl isobutylketone:isopropanol:methylethylketone (MIBK:IPA:MEK 1:3:1.5%) 2. Immerse the sample in the developer for 30-70 seconds 3. Rinse sample with IPA 4. Dry the sample with a Nitrogen gun PMMA SiO2 Silicon(100)

  33. AFM images Patterned PMMA

  34. 4. Growing Self-Assembled monolayer PMMA aminosilane Silicon(100) Immerse the patterned sample in 0.1 – 1.0 % aminosilane in water for 20-30 mins. Wash the sample with water Dry with the nitrogen gun

  35. AFM image PMMA trenches with aminosilane Trenches are still intact and no swelling

  36. Stability of siloxane bond during liftoff silicon silicon Siloxane bond ----Si----O-----Si---

  37. 5. Molecular Liftoff SiO2 aminsilane Silicon(100) Unexposed PMMA removal Dichloromethane (warm/hot) Acetone (room tempetature/warm) N-methyl-2-pyrrolidone (NMP) (room temperature)

  38. AFM images Bad liftoff (PMMA still left on silicon)

  39. AFM image Good liftoff

  40. 6. DNA origami attachment SiO2 DNA Silicon(100) Place few microliters DNA sample on the pattern Leave it for 1-2 hrs Wash the sample with water Dry the sample with nitrogen

  41. AFM image DNA origami on aminosilane anchor pads

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