Autonomous Bioluminescent Expression Of The Bacterial Luciferase Gene Cassette (Lux) In A Mammalian Cell Line, 2009 University of Tennessee - Knoxville
Autonomous Bioluminescent Expression Of The Bacterial Luciferase Gene Cassette (Lux) In A Mammalian Cell Line, Dan M. Close, Stacey S. Patterson, Steven Ripp, Seung J. Baek, John Sanseverino, Gary S. Sayler
Gary S. Sayler
The bacterial luciferase (lux) gene cassette consists of five genes (luxCDABE) whose protein products synergistically generate bioluminescent light signals exclusive of supplementary substrate additions or exogenous manipulations. Historically expressible only in prokaryotes, the lux operon was re-synthesized through a process of multi-bicistronic, codon-optimization to demonstrate for the first time self-directed bioluminescence emission in a mammalian HEK293 cell line in vitro and in vivo.
Autonomous in vitro light production was shown to be 12-fold greater than the observable background associated with untransfected control cells. The availability of reduced riboflavin phosphate (FMNH2) was identified as the limiting bioluminescence ...
The Construction And Analysis Of Marker Gene Libraries, 2009 University of Tennessee, Knoxville
The Construction And Analysis Of Marker Gene Libraries, S.M. Short, F. Chen, Steven Wilhelm
Marker genes for viruses are typically amplified from aquatic samples to determine whether specific viruses are present in the sample, or to examine the diversity of a group of related viruses. In this chapter, we will provide an overview of common methods used to amplify, clone, sequence, and analyze virus marker genes, and will focus our discussion on viruses infecting algae, bacteria, and heterotrophic flagellates. Within this chapter, we endeavor to highlight critical aspects and components of these methods. To this end, instead of providing a detailed experimental protocol for each of the steps involved in examining virus marker gene ...
Interference With Histidyl-Trna Synthetase By A Crispr Spacer Sequence As A Factor In The Evolution Of Pelobacter Carbinolicus, 2009 University of Massachusetts - Amherst
Interference With Histidyl-Trna Synthetase By A Crispr Spacer Sequence As A Factor In The Evolution Of Pelobacter Carbinolicus, Derek Lovley, Muktak Aklujkar
BACKGROUND: Pelobacter carbinolicus, a bacterium of the family Geobacteraceae, cannot reduce Fe(III) directly or produce electricity like its relatives. How P. carbinolicus evolved is an intriguing problem. The genome of P. carbinolicus contains clustered regularly interspaced short palindromic repeats (CRISPR) separated by unique spacer sequences, which recent studies have shown to produce RNA molecules that interfere with genes containing identical sequences. RESULTS: CRISPR spacer #1, which matches a sequence within hisS, the histidyl-tRNA synthetase gene of P. carbinolicus, was shown to be expressed. Phylogenetic analysis and genetics demonstrated that a gene paralogous to hisS in the genomes of Geobacteraceae ...
Synthetic Genome: Now That We’Re Creators, What Should We Create?, 2009 Wesleyan University
Synthetic Genome: Now That We’Re Creators, What Should We Create?, Frederick M. Cohan
Frederick M. Cohan
No abstract provided.
The Ecology Of Speciation In Bacillus, 2009 Wesleyan University
The Ecology Of Speciation In Bacillus, Nora Connor, Johannes Sikorski, Alejandro P. Rooney, Sarah Kopac, Alexander F. Koeppel, Andrew Burger, Scott G. Cole, Elizabeth B. Perry, Danny Krizanc, Nicholas C. Field, Michele Slaton, Frederick M. Cohan
Frederick M. Cohan
No abstract provided.
The Genome Of Geobacter Bemidjiensis, Exemplar For The Subsurface Clade Of Geobacter Species That Predominate In Fe(Iii)-Reducing Subsurface Enviorments, 2009 University of Massachusetts - Amherst
The Genome Of Geobacter Bemidjiensis, Exemplar For The Subsurface Clade Of Geobacter Species That Predominate In Fe(Iii)-Reducing Subsurface Enviorments, Derek Lovley, Muktak Aklujkar, Nealson D. Young, Dawn Holmes, Milind Chavan, Carla Risso, Hajnalka E. Kiss, Cliff S. Han, Miriam L. Land
BACKGROUND: Geobacter species in a phylogenetic cluster known as subsurface clade 1 are often the predominant microorganisms in subsurface environments in which Fe(III) reduction is the primary electron-accepting process. Geobacter bemidjiensis, a member of this clade, was isolated from hydrocarbon-contaminated subsurface sediments in Bemidji, Minnesota, and is closely related to Geobacter species found to be abundant at other subsurface sites. This study examines whether there are significant differences in the metabolism and physiology of G. bemidjiensis compared to non-subsurface Geobacter species. RESULTS: Annotation of the genome sequence of G. bemidjiensis indicates several differences in metabolism compared to previously sequenced ...
Regulatory Elements Of Xenopus Col2a1 Drive Cartilaginous Gene Expression In Transgenic Frogs, 2009 Dalhousie University
Regulatory Elements Of Xenopus Col2a1 Drive Cartilaginous Gene Expression In Transgenic Frogs, Ryan Kerney, Brian K. Hall, James Hanken
This study characterizes regulatory elements of collagen 2α1 (col2a1) in Xenopus that enable transgene expression in cartilage-forming chondrocytes. The reporters described in this study drive strong cartilage-specific gene expression, which will be a valuable tool for further investigations of Xenopus skeletal development. While endogenous col2a1 mRNA is expressed in many embryonic tissues, its expression becomes restricted to tadpole and adult chondrocytes. This chondrocyte-specific expression is recapitulated by col2a1 reporter constructs, which were tested through I-SceI meganuclease-mediated transgenesis. These constructs contain a portion of the Xenopus tropicalis col2a1 intron, which aligns to a cartilage-specific intronic enhancer that has been well characterized ...
Quantitativepcrmethods Forrna Anddnainmarine Sediments: Maximizing Yieldwhile Overcoming Inhibition, Karen Lloyd, Barbara J. Macgregor, Andreas Teske
For accurate quantification of DNA and RNA from environmental samples, yield loss during nucleic acid purification has to be minimized. Quantitative PCR (qPCR) and reverse transcription (RT)-qPCR require a trade-off between maximizing yield and removing inhibitors. We compared DNA and RNA yield and suitability for quantitative SYBR Green PCR and RT-PCR using the UltraClean and PowerSoil extraction kits and a bead-beating protocol with phenol/chloroform extraction steps. Purification methods included silica-column-based procedures from the MoBio kits, RNeasy MinElute, WizardPlus miniprep columns, and an acrylamide gel extraction. DNA and RNA purification with WizardPlus and RNeasy, respectively, led to significant losses ...
Determining Rates Of Virus Production In Aquatic Systems By The Virus Reduction Approach,, 2009 University of Tennessee, Knoxville
Determining Rates Of Virus Production In Aquatic Systems By The Virus Reduction Approach,, M.G. Weinbauer, J.M. Rowe, Steven Wilhelm
The reduction approach to assess virus production and the prokaryotic mortality by viral lysis stops new infection by reducing total virus abundance (and thus virus–host contacts). This allows for easy enumeration of viruses that originate from lysis of already infected cells due to the decreased abundance of free virus particles. This reoccurrence can be quantified and used to assess production and cell lysis rates. Several modifications of the method are presented and compared. The approaches have great potential for elucidating trends in virus production rates as well as for making generalized estimates of the quantitative effects of viruses on ...
The Manual Of Aquatic Virus Ecology, 2009 University of Tennessee, Knoxville
The Manual Of Aquatic Virus Ecology, Steven Wilhelm
No abstract provided.