A Synthetic Electronic Nanopore for DNA

Sequencing and Stochastic Sensing

Mr. Aaron Choi, Computer Science, Sophomore

Mr. Davis Sneider, Biomedical Engineering, Sophomore

Mr. Saifuddin Aijaz, Chemical Engineering, Pre-Junior

Mentors:

Dr. David Wendell, Assistant Professor, Environmental Engineering

Dr. Vasile Nistor, Assistant Professor, Biomedical Engineering

Ms. Elizabeth Wurtzler, Graduate Student, Environmental Engineering

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Introduction

• Background

• Goals & Tasks

• Time Schedule– What we’ve done, where we’re going

• Inserting DNA– What we’re looking for, what we’ve found

• Findings

• Conclusion

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Current problem

• DNA sequencing can cost up to 10,000 dollars and take about a week

• Nanopore technology can save a lot of money and reduce the time to one day

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Nanopores: What are they?

• They are extremely small holes.

• They have potential applications for many kinds of developing technology

Oxford Nanopore Technologies

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Hydraphile Nanopore

• A synthetic nanopore, created by Dr. George Gokel at University of Missouri, St. Louis

• Lariat Ethers– Excellent cation selectivity– Excellent binding and release kinetics

Royal Society of Chemistry http://pubs.rsc.org/en/content/articlehtml/2000/cc/a903825f

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Size Comparison• The nanopore is said to be

approximately 8 picometers

• DNA has been shown to go through the nanopore and single stranded DNA is 1 nm

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Applications

• We could detect cancer earlier and much more efficiently

• DNA sequencing allows us to find many genetic disorders

• Ability to detect viruses

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Our Goals

• To determine which buffer works best

• To use the hydraphile nanopore for– DNA sequencing– Norovirus sensing

• Help to define the width of the hydraphile nanopore.

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Tasks• Use QuB to analyze data from four buffers

• Run items through nanopore– Ion Solutions– DNA – Norovirus

• Use passages to get an idea of how wide the nanopore is

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Time Schedule

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Conclusion From Buffers Tests

• Out of the four solutions used, it was determined that KCl is the best choice to use for nanopore sequencing as it gives a more stable membrane.

apcg.space.noa.gr

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Potassium Buffer

• 1M KCl Buffer, with 5mM Hepes

• Able to get data with ease

• Analyzing Data– Clampex

• 100< data points• Standard deviation

1.76 nanosiemens Glogster.com

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Painting the Membrane

• Take in and remove lipid hexane solution

• Create air bubble with pipet

• Wipe air bubble over membrane

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Inserting Nanopores

• Once a thin membrane is present, we then insert the hydraphile nanopore

• If membrane is too thick, nanopores won’t span length of membrane

Wikipedia http://en.wikipedia.org/wiki/Synthetic_ion_channels

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Nanopore Insertion

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Inserting DNA

• Dilute mixture– 2µL of DNA– 18µL of water

• Intake .5µL of mixture overtop of hole

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Detecting DNA Current Change

• Inserting DNA causes resistances in the current across the membrane– Negative charge across membrane

www.ks.uiuc.edu

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DNA Passing

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Resistance Zoomed19

What We Measured

• 2 major measurements– Blockage %– Dwell Time (ms)

• DNA length– 250 BP– 500 BP– 1,000 BP– 2,500 BP

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What Does It Mean & What Is It Useful For?

• Blockage %– Tells us how much of the nanopore has been blocked

– Helps us identify approximate width of DNA/RNA strand

• Event Duration– Tells us how long it took the DNA

segment to pass through the nanopore

– Helps us identify approximate length of the DNA/RNA strand

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References

• Gokel, George. Hydraphiles: Design, Synthesis and Analysis of a Family of Synthetic, Cation-conducting Channels. Tech. Royal Society of Chemistry, 24 Dec. 1999. Web. 13 June 2014.

• "Towards the 15-minute Genome." The Economist. The Economist Newspaper, 12 Mar. 2011. Web. 17 June 2014.

• Uddin A, Yemenicioglu S, Chen C-H, Corigliano E, Milaninia K and Theogarajan L. Integration of solid-state nanopores in a 0.5 um CMOS foundry process. Nanotechnology. IOPScience, 31 October 2013. Web. 2 July 2014.

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Thank You!

• We would like to thank NSF for funding our research [Grant ID No.: DUE – 0756921]

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Questions?

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