Erie Group Zimeng Li 2012-5-30 AFM real time visualization (7.01) A Fluid/Video AFM study of MMR...
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Erie Group Zimeng Li 2012-5-30 AFM real time visualization (7.01) A Fluid/Video AFM study of MMR (2.29) AFM visualization of MutS Sliding Clamp Formation in DNA Mismatch Repair (5.30) ? Observation of Protein-DNA Interactions in real time (fall) Distinguish Proteins/Protein-DNA complex/EFM study Observation of Protein- DNA interaction involved in MMR Fluorescence-AFM Hybrid Imaging Distinguish Proteins/Protein-DNA complex/FRET study Ultramicroscopy AFM visualization of MutS Sliding Clamp Formation in DNA Mismatch Repair
Erie Group Zimeng Li 2012-5-30 AFM real time visualization (7.01) A Fluid/Video AFM study of MMR (2.29) AFM visualization of MutS Sliding Clamp Formation
Erie Group Zimeng Li 2012-5-30 AFM real time visualization
(7.01) A Fluid/Video AFM study of MMR (2.29) AFM visualization of
MutS Sliding Clamp Formation in DNA Mismatch Repair (5.30) ?
Observation of Protein-DNA Interactions in real time (fall)
Distinguish Proteins/Protein-DNA complex/EFM study Observation of
Protein-DNA interaction involved in MMR Fluorescence-AFM Hybrid
Imaging Distinguish Proteins/Protein-DNA complex/FRET study
Ultramicroscopy AFM visualization of MutS Sliding Clamp Formation
in DNA Mismatch Repair
Slide 2
Contents DNA Mismatch Repair Sliding Clamp Formation AFM
techniques in fluid Results Future To-dos
Slide 3
DNA Mismatch Repair
Slide 4
Sliding Clamp Formation Tessmer, et.al (2008)
Slide 5
Sliding Clamp Formation Qiu, et.al (2012)
Slide 6
Myosin V walk/Nano robot spider walk Super resolution
microscopy (Yildiz(2003);Nils Walter) AFM (Kodera(2010);Nils
Walter) FRET (Qiu(2012)) ? Techniques
Slide 7
Atomic Force Microscopy
Slide 8
From Force to Distance
Slide 9
AFM
Slide 10
Slide 11
Fluid AFM techniques Difference between thermal tune and
cantilever tune Substrates, special tips/treatment Atomic Force
Microscopy
Slide 12
Results
Slide 13
Recall
Slide 14
DNA Sample Buffer APTES?Rinse?In Air? Image Buffer Tip
WaterLoSHiSNoS DryFluidDryFluidDryFluidDryFluid Decent Image29%
(6/21) 18% (2/11) 50% (1/2) 100% (1/1) Great Image38% (8/21) 18%
(2/11) 13% (1/8) Total67%36%50%13%0%(1)100% Success Probability Tip
consumption: ~70, 30% gets DNA, 30% gets something real, 30%
bad
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Great imageDecent image New tips31 Old tips12 My operations
wrong?
Slide 46
15min Running a full cleaning cycle Deposition/No incubation
Rinse once (Twice results in no DNA?) 15-30min/cycle Image/No
Good?/Change tips/Image/ Average 2~3tips/sample Current
Protocol
Slide 47
Continue working with lo-salt buffer If not working, try
no-salt buffer Adding NaCl to increase DNA mobility Hi-salt using
APTES treated mica to reduce mobility Does MutS work in water or
no-salt buffer? To-dos