Lyle R. Middendorf

LI-COR Biosciences (United States)

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Stephen C. Roemer

LI-COR Biosciences (United States)

Narasimhachari Narayanan

LI-COR Biosciences (United States)

Robert Bruce

Johns Hopkins University Applied Physics Laboratory

John Brumbaugh

University of Nebraska–Lincoln

Patrick G. Humphrey

LI-COR Biosciences (United States)

Rainer Erdmann

PicoQuant (Germany)

Steven A. Soper

University of Kansas

Robert D. Eckles

LI-COR Biosciences (United States)

Dan Grone

LI-COR Biosciences (United States)

Rex A. Peterson

LI-COR Biosciences (United States)

Cooperative Institutions

LI-COR Biosciences (United States)

University of Nebraska–Lincoln

Stanford University

Louisiana State University

University of Maryland, Baltimore

Pennsylvania State University

University of Michigan–Ann Arbor

Stanford Medicine

Georgia State University

University of Minnesota

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Automated bead-trapping apparatus and control system for single-molecule DNA sequencing

Optics Express (2008)

Greg Bashford Don C. Lamb Dan Grone Bob Eckles K. Kornelsen

We have been investigating a microfluidics platform for high-speed, low-cost sequencing of single DNA molecules using novel "charge-switch" nucleotides. A significant challenge is the design of a flowcell suitable for manipulating bead-DNA complexes and sorting labeled polyphosphate molecules by charge. The flowcell is part of a single-molecule detection instrument, creating fluorescence images from labeled polyphosphates. These images would ultimately be analyzed by signal processing algorithms to identify specific nucleotides in a DNA sequence. Here we describe requirements of the fluidics system for loading, identifying, tracking, and positioning beads. By dynamically modulating pressure gradients in the plenum chambers of a multi-channel network, we could guide individual beads with high precision to any desired coordinate and reversibly trap them in stepped channels. We show that DNA immobilized on pressure-trapped beads can be physically extended into a downstream channel under electric force for analysis. Custom dynamic algorithms for automated bead control are described.

Fluidics

Bead

Cartridge

SIGNAL (programming language)

10.1364/oe.16.003445

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Citations (7)

Analysis of tyrosinase gene mutations using direct automated infrared fluorescence DNA sequencing of amplified exons

Electrophoresis (1994)

William S. Oetting James P. Fryer Yasuhisa Oofuji Lyle R. Middendorf John Brumbaugh

Abstract The ability to correctly diagnose the molecular cause of genetic diseases is becoming increasingly important in medicine. This requires an efficient method for the analysis of the DNA sequence of specific genes and the detection of mutations in affected individuals. We report a method to determine the mutations responsible for tyrosinase related albinism (OCA1) using a combination of polymerase chain reaction‐single stranded conformational polymorphism (PCR‐SSCP) analysis and direct DNA cycle sequencing using fluorescently labeled oligonucleotides and an automated DNA sequencer based on infrared fluorescence technology. This method allows DNA from several individuals to be sequenced quickly and simultaneously so that the specific location of each mutation and the carrier status of family members can be determined.

Single-strand conformation polymorphism

10.1002/elps.1150150127

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Citations (25)

<title>Two-dimensional infrared fluorescence scanner used for DNA analysis</title>

Proceedings of SPIE, the International Society for Optical Engineering/Proceedings of SPIE (1995)

Lyle R. Middendorf Robert Bruce John Brumbaugh Daniel L. Grone Gi Young Jang

A LI-COR Model 4000 DNA Sequencer has been modified by removing the internal scanning infrared fluorescence microscope and combining it with an external, orthogonal scanner. Due to the reduced background fluorescence and light scattering of nylon membranes in the near- infrared (8000 nm) as compared to the visible region of the optical spectrum, sensitivity of labeled DNA fragments is enhanced. Dot blots of dilution series of labeled oligonucleotides reveal a detection limit of 25 attomole (25 X 10^-18 mole). DNA fragments blotted onto nylon membranes using direct transfer electrophoresis in multiplex DNA sequencing can also be detected and subsequently analyzed.

10.1117/12.208522

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Citations (6)

Correlation between Scattering Properties of Silver Particle Arrays and Fluorescence Enhancement

Applied Spectroscopy (2008)

Henryk Szmacinski Joseph R. Lakowicz Jeffrey M. Catchmark Khalid Eid Jon P. Erson

We report on the nanofabrication of patterned silver particle arrays using electron-beam lithography and the evaluation of their optical properties using backscattering and fluorescence spectroscopy. The silver particles varied in size from 100 to 250 nm and were in the shape of circles, squares, and triangles. Three inter-particle separations, 40, 65, and 90 nm as measured from the side of one particle to the side of the next particle, were used. We observed distinctive patterns of backscattering and fluorescence intensity depending on the particle size, inter-particle spacing, and excitation/emission wavelength used. Our approach allows for a study of the correlation between the backscattering intensities and fluorescence enhancement of silver particle arrays, which can be used to optimize the arrays for multi-fluorophore configuration for advanced sensing designs.

Particle (ecology)

Fluorescence Correlation Spectroscopy

10.1366/000370208784909553

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Citations (34)

Infrared Fluorescent Detection of D1S80 Alleles from Blood and Body Fluid Collected on IsoCode™ Devices

BioTechniques (1997)

Reena Roy Lyle R. Middendorf

A genetic locus D1S80 (pMCT 118) containing variable number of tandem repeats (VNTR) has been used extensively in forensic analysis and paternity testing. In the current research, DNA was isolated from blood saliva and nasal secretions collected on two types of IsoCode paper-based devices. The D1S80 locus was amplified using PCR technology, and the alleles were separated by gel electrophoresis and then detected using an infrared (IR) fluorescence automated DNA sequencer. IR-labeled amplification products were generated from human genomic DNA using oligonucleotide primers, which were covalently linked to an infrared fluorescent dye (IRD41) at the 5' end. This system combines IR fluorescence chemistry and laser technology, thus eliminating the need for post-electrophoretic gel handling for the detection of the alleles. Real-time detection after separation of the alleles is valuable for visualization of the data. The VNTR alleles are displayed as familiar autoradiogram-like images, which can also be analyzed by computer. Since DNA is eluted from the IsoCode devices only with sterile distilled water and without time-consuming methods of extraction, amplification can be performed from numerous samples within a short period of time.

genomic DNA

Body fluid

10.2144/97235pf02

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Citations (12)

Continuous, On-Line, Real Time DNA Sequencing Using Multifluorescently Tagged Primers

Springer eBooks (1991)

John Brumbaugh Lyle R. Middendorf Dan Grone Jerry L. Ruth

Sequence (biology)

10.1007/978-1-4757-9513-4_3

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Citations (0)

Time-resolved photon counting for multiplexed DNA electrophoreses separations

Lloyd M. Davis Jim Amen Dan Draney Lyle R. Middendorf Rex A. Peterson

Monte Carlo simulations are used to evaluate time resolved photon counting as a means for identifying and quantifying overlapping electropherogram bands, which have been labeled by dyes with disparate fluorescence lifetimes.

Photon Counting

Electropherogram

10.1364/lacea.2000.fa4

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Citations (0)

Enhanced throughput for infrared automated DNA sequencing

Proceedings of SPIE, the International Society for Optical Engineering/Proceedings of SPIE (1995)

Lyle R. Middendorf Bill O. Gartside Pat G. Humphrey Stephen C. Roemer David Sörensen

Several enhancements have been developed and applied to infrared automated DNA sequencing resulting in significantly higher throughput. A 41 cm sequencing gel (31 cm well- to-read distance) combines high resolution of DNA sequencing fragments with optimized run times yielding two runs per day of 500 bases per sample. A 66 cm sequencing gel (56 cm well-to-read distance) produces sequence read lengths of up to 1000 bases for ds and ss templates using either T7 polymerase or cycle-sequencing protocols. Using a multichannel syringe to load 64 lanes allows 16 samples (compatible with 96-well format) to be visualized for each run. The 41 cm gel configuration allows 16,000 bases per day (16 samples X 500 bases/sample X 2 ten hour runs/day) to be sequenced with the advantages of infrared technology. Enhancements to internal labeling techniques using an infrared-labeled dATP molecule (Boehringer Mannheim GmbH, Penzberg, Germany) and Sequenase (U.S. Biochemical) have also been made. The inclusion of glycerol in the sequencing reactions yields greatly improved results for some primer and template combinations. The inclusion of (alpha) -Thio-dNTP's in the labeling reaction increases signal intensity two- to three-fold.

Primer (cosmetics)

Template

10.1117/12.206009

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Citations (7)

DNA Technologies: Sequencing Technology

Lyle R. Middendorf Patrick G. Humphrey Narasimhachari Narayanan Stephen C. Roemer

This chapter contains sections titled: Introduction Overview of Sanger Dideoxy Sequencing Fluorescence Dye Chemistry Biochemistry of DNA Sequencing Fluorescence DNA Sequencing Instrumentation DNA Sequence Analysis References

10.1002/9783527612642.ch07

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A Programmable Graphics Processor based on Partial Stream Rewriting

Computer Graphics Forum (2013)

Lyle R. Middendorf Christian Haubelt

Abstract Current graphics processing units (GPU) typically offer only a limited number of programmable pipeline stages, whose usage, data flow and topology are mostly fixed. Although a more flexible, custom rendering pipeline can be emulated using the compute functionality of existing GPUs, this approach requires to manage work queues, synchronization, and scheduling in software. In this paper, we present a hardware architecture for a novel, programmable rendering pipeline, which is based on a circulating stream of data and control tokens that are iteratively modified via pattern matching. Our architecture provides light‐weight mechanisms for dynamic thread creation, lock‐free synchronization, and scheduling to support recursion, dynamic shader linkage and custom primitive types. A hardware prototype, running complex examples, demonstrates the improved reconfigurability also the scalability of our graphics architecture.

Shader

Stream Processing

Parallel rendering

10.1111/cgf.12240

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Citations (5)