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University of California Davis

17 ARTICLES PUBLISHED IN JoVE

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Neuroscience

Assaying Locomotor Activity to Study Circadian Rhythms and Sleep Parameters in Drosophila
Joanna C. Chiu 1,2, Kwang Huei Low 1,3, Douglas H. Pike 1, Evrim Yildirim 1,3, Isaac Edery 1,3
1Center for Advanced Biotechnology and Medicine, Rutgers University, 2Current Address: Department of Entomology, College of Agricultural and Environmental Sciences, University of California, Davis, 3Department of Molecular Biology and Biochemistry, Rutgers University

We describe procedures for recording daily locomotor activity rhythms of Drosophila and subsequent data analysis. Locomotor activity rhythms are a reliable behavioral output of animal circadian clocks and are used as the standard readout of clock function when studying circadian mutants or examining how the environment regulates the circadian system.

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Bioengineering

Low-Cost Cryo-Light Microscopy Stage Fabrication for Correlated Light/Electron Microscopy
David B. Carlson 1, James E. Evans 1
1Department of Molecular and Cellular Biology, University of California Davis

We demonstrate the fabrication of a low-cost cryogenic stage designed to fit most reflected light microscopes. This lab-built cryogenic stage enables efficient and reliable correlative imaging between cryo-light and cryo-electron microscopy.

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Bioengineering

On-Chip Endothelial Inflammatory Phenotyping
J. Sherrod DeVerse *1, Keith A. Bailey *1, Greg A. Foster 1, Vaishali Mittal 1, Stuart M. Altman 1, Scott I. Simon 1, Anthony G. Passerini 1
1Department of Biomedical Engineering, University of California, Davis

Microfluidic flow chambers etched by photolithography and fabricated from PDMS are applied to probe functional outcomes associated with EC dysfunction and inflammation. In a representative experiment, the ability of differential shear stress to modulate monocytic cell adhesion to cytokine activated EC monolayers is demonstrated.

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Biology

LeafJ: An ImageJ Plugin for Semi-automated Leaf Shape Measurement
Julin N. Maloof *1, Kazunari Nozue *1, Maxwell R. Mumbach 1, Christine M. Palmer 1
1Department of Plant Biology, University of California Davis

Demonstration of key methods for high throughput leaf measurements. These methods can be used to accelerate leaf phenotyping when studying many plant mutants or otherwise screening plants by leaf phenotype.

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Biology

Optical Mapping of Intra-Sarcoplasmic Reticulum Ca2+ and Transmembrane Potential in the Langendorff-perfused Rabbit Heart
Lianguo Wang 1, Nicole M. De Jesus 2, Crystal M. Ripplinger 1
1Department of Pharmacology, University of California Davis, 2Departments of Pharmacology and Biomedical Engineering, University of California Davis

This article describes the detailed protocol and equipment necessary for dual optical mapping of transmembrane potential (Vm) and free intra-sarcoplasmic reticulum (SR) Ca2+ in the Langendorff-perfused rabbit heart. This method allows for direct observation and quantification of Vm and SR Ca2+ dynamics in the intact heart.

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Biology

Microperfusion Technique to Investigate Regulation of Microvessel Permeability in Rat Mesentery
Fitz-Roy E. Curry 1, Joyce F. Clark 1, Roger H. Adamson 1
1Department of Physiology & Membrane Biology, University of California Davis

The modified Landis technique enables paired measurement of the hydraulic conductivity of individual microvessels in the mesentery of normal and genetically modified rats under control and test conditions using microperfusion techniques. It provides a convenient method to evaluate mechanisms that regulate microvessel permeability and transvascular exchange under physiological conditions.

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Bioengineering

Surface Functionalization of Hepatitis E Virus Nanoparticles Using Chemical Conjugation Methods
Chun Chieh Chen *1, Marie Stark *1, Mo Baikoghli 1, R. Holland Cheng 1
1Department of Molecular and Cellular Biology, University of California Davis

We have engineered the capsid protein of hepatitis E virus as a theranostic nanoparticle (HEVNP). HEVNP self-assembles into a stable icosahedral cage in mucosal delivery. Here, we describe the modification of HEVNPs for tumor targeting by mutating surface-exposed residues to cysteines, which conjugate synthetic ligands that specifically bind tumor cells.

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Environment

Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions
Andrew J. Margenot 1, Sanjai J. Parikh 2, Francisco J. Calderón 3
1Department of Crop Sciences, University of Illinois Urbana-Champaign, 2Department of Land, Air and Water Resources, University of California Davis, 3Central Great Plains Resources Management Research, USDA ARS

SOM underlies many soil functions and processes, but its characterization by FTIR spectroscopy is often challenged by mineral interferences. The described method can increase the utility of SOM analysis by FTIR spectroscopy by subtracting mineral interferences in soil spectra using empirically obtained mineral reference spectra.

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Medicine

The Perinatal Asphyxiated Lamb Model: A Model for Newborn Resuscitation
Payam Vali *1, Sylvia Gugino *2, Carmon Koenigsknecht 2, Justin Helman 2, Praveen Chandrasekharan 2, Munmun Rawat 2, Satyan Lakshminrusimha 1, Jayasree Nair 2
1Department of Pediatrics, University of California Davis, 2Department of Pediatrics, University at Buffalo

Invasive instrumentation of the fetal lamb provides accurate physiologic measurements of the transitioning circulation in a model that closely mimics the newly born infant.

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Biochemistry

Measuring Liver Mitochondrial Oxygen Consumption and Proton Leak Kinetics to Estimate Mitochondrial Respiration in Holstein Dairy Cattle
Heidi A. Rossow 1, Gabriela Acetoze 2, John Champagne 1, Jon J. Ramsey 3
1Veterinary Medicine Teaching and Research Center, University of California Davis, 2Archer Daniels Midland, 3Department of Molecular Biosciences, University of California Davis

Here, we share methods for measuring mitochondrial oxygen consumption, a defining concept of nutritional energetics, and proton leak, the primary cause of inefficiency in mitochondrial generation of ATP. These results can account for 30% of the energy lost in nutrient utilization to help evaluate mitochondrial function.

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Immunology and Infection

A Mouse Model to Assess Innate Immune Response to Staphylococcus aureus Infection
Leif S. Anderson 1, Mack B. Reynolds 1, Kathryn R. Rivara 1, Lloyd S. Miller 2, Scott I. Simon 1
1Department of Biomedical Engineering, University of California Davis, 2Department of Dermatology, Johns Hopkins University School of Medicine

An approach is described for real-time detection of the innate immune response to cutaneous wounding and Staphylococcus aureus infection of mice. By comparing LysM-EGFP mice (which possess fluorescent neutrophils) with a LysM-EGFP crossbred immunodeficient mouse strain, we advance our understanding of infection and the development of approaches to combat infection.

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Immunology and Infection

Isolation and Quantification of Zika Virus from Multiple Organs in a Mouse
James D. Brien *1, Mariah Hassert *1, E. Taylor Stone 1, Elizabeth Geerling 1, Lillian Cruz-Orengo 2, Amelia K. Pinto 1
1Department of Molecular Microbiology and Immunology, Saint Louis University, 2School of Veterinary Medicine University of California Davis

The goal of the protocol is to demonstrate the techniques used to investigate viral disease by isolating and quantifying Zika virus, from multiple organs in a mouse following infection.

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Biochemistry

A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Katherine M. Murphy 1, Siwon Chung 1, Shruti Fogla 1, Hana B. Minsky 1, Karen Yong Zhu 1, Philipp Zerbe 1
1Department of Plant Biology, University of California Davis

Here we present easy to use protocols for producing and purifying diterpenoid metabolites through the combinatorial expression of biosynthetic enzymes in Escherichia coli or Nicotiana benthamiana, followed by chromatographic product purification. The resulting metabolites are suitable for various studies including molecular structure characterization, enzyme functional studies, and bioactivity assays.

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Neuroscience

Reliable Isolation of Central Nervous System Microvessels Across Five Vertebrate Groups
Yinyu Yuan *1, Jacquelyn R. Dayton *1, Marie-Lena Freese 2, Bryce G. Dorflinger 1, Lillian Cruz-Orengo 1
1Department of Anatomy, Physiology and Cell Biology, University of California Davis, 2University of Veterinary Medicine Hannover Foundation

The goal of this protocol is to isolate microvessels from multiple regions of the central nervous system of lissencephalic and gyrencephalic vertebrates.

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Biology

A Magnetic-Bead-Based Mosquito DNA Extraction Protocol for Next-Generation Sequencing
Tse-Yu Chen 1, Adam E. Vorsino 2, Kyle J. Kosinski 1, Ana L. Romero-Weaver 1, Eva A. Buckner 1, Joanna C. Chiu 3, Yoosook Lee 1
1Florida Medical Entomology Laboratory, Department of Entomology and Nematology, Institute of Food and Agricultural Sciences, University of Florida, 2U. S. Fish and Wildlife Service, Pacific Islands Fish and Wildlife Office, 3Department of Entomology and Nematology, University of California Davis

Described here is a DNA extraction protocol using magnetic beads to produce high quality DNA extractions from mosquitoes. These extractions are suitable for a downstream next-generation sequencing approach.

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Immunology and Infection

Cell-Free Scaled Production and Adjuvant Addition to a Recombinant Major Outer Membrane Protein from Chlamydia muridarum for Vaccine Development
Sean F. Gilmore *1, Wei He *1, Angela C. Evans 1, Delia F. Tifrea 2, Sukumar Pal 2, Brent Segelke 1, Sandra K. G. Peters 1, B. Dillon Vannest 1, Nicholas O. Fischer 1, Amy Rasley 1, Luis M. de la Maza 2, Matthew A. Coleman 1,3
1Physical and Life Sciences Directorate, Lawrence Livermore National Laboratory, 2Department of Pathology and Laboratory Medicine, University of California, 3School of Medicine, Radiation Oncology, University of California Davis

This protocol describes using commercial, cell-free protein expression kits to produce membrane proteins supported in nanodisc that can be used as antigens in subunit vaccines.

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Biology

Isolation of Small Preantral Follicles from the Bovine Ovary Using a Combination of Fragmentation, Homogenization, and Serial Filtration
Stephanie P. McDonnell 1, Juliana I. Candelaria 1, Amanda J. Morton 1, Anna C. Denicol 1
1Department of Animal Science, University of California Davis

Advancing the study of preantral folliculogenesis requires efficient methods of follicle isolation from single ovaries. Presented here is a streamlined, mechanical protocol for follicle isolation from bovine ovaries using a tissue chopper and homogenizer. This method allows collection of a large number of viable preantral follicles from a single ovary.

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