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Methods and Compositions for Selective Testing of Mammalian Proteomics From Mixed Biological Environments

96 Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;} Studies of heterochronic parabiosis (surgical joining of young and old animals) suggest both productive tissue repair and the key signal transduction pathways that control stem cell activation are restored to ‘youth’ in the old parabionts by young systemic factors.  It would be beneficial from academic and clinical stand-points to determine which proteins in tissues of parabiotically connected animals are derived from the circulation of young versus old partner. Such a database of systemic proteins that end up in specific tissues would suggest potentially rejuvenating (young blood) and inhibitory (old blood) molecules with direct effects in a given tissue. While biochemical fractionation of serum and plasma can provide some characterization of the molecular differences between young and old circulatory milieu, this technique is fraught with the risk of missing proteins that act in complexes with each other and other macromolecules. UC Berkeley researchers have discovered methods and compositions that overcome these problems by relying on tRNA synthase that specifically recognizes and incorporates Bio-Orthogonal Non-Canonical Amino acid Tagging (“BONCAT”) into proteins. To facilitate detection by proteomics, we have selected the BONCAT method over the cell type-specific labeling with amino acid precursors where proteomes are tagged with heavy isotope—labeled precursors; and over the incorporation of Met analogs azidohomoalanine and homopropargylglycine, which do not allow one to selectively profile young versus old proteomes in settings of parabiosis. The researchers have also developed a novel transgenic mouse strain which demonstrate the survival and vigor of these animals as well as the effective proteome labeling of cells in vitro and all examined tissues in vivo.

Gene Delivery Into Mature Plants Using Carbon Nanotubes

96 Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;} Current methods of biomolecule delivery to mature plants are limited due to the presence of plant cell wall, and are additionally hampered by low transfection efficiency, high toxicity of the transfection material, and host range limitation. For this reason, transfection is often limited to protoplast cultures where the cell wall is removed, and not to the mature whole plant.  Unfortunately, protoplasts are not able to regenerate into fertile plants, causing these methods to have low practical applicability. Researchers at the University of California have developed a method for delivery of genetic materials into mature plant cells within a fully-developed mature plant leaf, that is species-independent. This method utilizes a nano-sized delivery vehicle for targeted and passive transport of biomolecules into mature plants of any plant species. The delivery method is inexpensive, easy, and robust, and can transfer biomolecules into all phenotypes of any plant species with high efficiency and low toxicity.

Improved Energy Harvesting for Current-Carrying Conductors

There are an estimated 130 million wooden poles that support overhead power lines in the US.  Extreme weather, aging, storms or sabotage can all lead to potential damage of these poles and power lines, which can leave large areas without basic necessities.  Due to this risk, it’s anticipated that power utility companies will deploy sensors and corresponding energy harvesters to better respond to potential damage of this critical electricity grid infrastructure. To address this anticipated mass deployment of sensors and harvesters, researchers at UC Berkeley have developed technology improvements to harvesting of electrical energy from energized conductors carrying alternating currents, such as those on overhead and underground power lines (as well as power-supplying conductors in offices and dwellings).  These enhanced harvesters would improve the economics of deploying sensors across a national power grid.  The Berkeley harvesters can readily provide enough power to supply wireless communication devices, energy storage batteries and capacitors, as well as sensors such as accelerometers, particulate matter measuring devices, and atmospheric sensors.


96 Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:Calibri; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;} Citrus pulp and sugar beet pulp are pectin-rich agricultural wastes that are globally produced in significant amounts and have the potential to contribute towards the greater bioeconomy as a source of raw, inexpensive carbohydrate biomass. There is currently limited use for these waste streams. In some cases, pulps are dried, pelleted, and repurposed as an inexpensive livestock feed, however this application is barely profitable due to high production costs. There is a need for technologies that can cost-effectively transform pectin-rich waste streams into value-added products of commercial interest.   UC Berkeley researchers developed an efficient microbial strain technology and metabolic fermentation methods for the bioconversion of pectin-rich waste streams to useful bio-based commodity chemicals and biofuels. In addition to the beneficial environmental impact of utilizing a waste-stream, the fermentation technologies achieve three design goals set to optimize the productivity of bioconversions and economic viability. First, the technology allows for anaerobic fermentation, eliminating the need for culture oxygenation. This lowers operating costs by simplifying the metabolic requirements of high-density fermentation cultures. Second, co- utilization of the major component monosaccharides in the hydrolysate broth allows for productive conversion of the predominant, energy- rich biomass sugars. Third, fermentations can be conducted at low pH, discouraging contaminant growth and eliminating the need to buffer the hydrolysate mixture.  

Precision Irrigation System Using Passive Mechanical Valves And Mobile Robots

Prolonged drought in California and the Southwest has both severely reduced water allocation to farmers, and substantially increased water prices. As the drought continues, so does the pressure to increase water use efficiency and streamline water delivery practices in agriculture. The systems currently in use are insufficiently precise to satisfy the demands of high value crops such as almonds and grapes, which often require watering regimes tailored to individual plants.UC Berkeley researchers have developed a low-cost system of mechanical valves and mobile robots that will address this issue. One or more valves can be installed per plant, and periodically adjusted by the robots based on sensor data. The system provides a fine-grained control of water flow to compensate for factors that vary across the planting region.

An Ultra-Sensitive Method for Detecting Molecules

To-date, plasmon detection methods have been utilized in the life sciences, electrochemistry, chemical vapor detection, and food safety. While passive surface plasmon resonators have lead to high-sensitivity detection in real time without further contaminating the environment with labels. Unfortunately, because these systems are passively excited, they are intrinsically limited by a loss of metal, which leads to decreased sensitivity. Researchers at the University of California, Berkeley have developed a novel method to detect distinct molecules in air under normal conditions to achieve sub-parts per billion detection limits, the lowest limit reported. This device can be used detecting a wide array of molecules including explosives or bio molecular diagnostics utilizing the first instance of active plasmon sensor, free of metal losses and operating deep below the diffraction limit for visible light.  This novel detection method has been shown to have superior performance than monitoring the wavelength shift, which is widely used in passive surface plasmon sensors. 

Identification Of Sites For Internal Insertions Into Cas9

The ability to add a protein domain of new function is a standard molecular biology technique, and usually the domain is fused to a protein terminus. The CRISPR-associated protein Cas9 already has widespread utility for genome engineering, yet adding protein domains would increase precision and specificity. Both protein termini of Cas9, however, are close to each other and in a small defined region, which limits the effectiveness of standard fusion approaches. Therefore, insertion sites within Cas9 that will not disrupt Cas9 function are needed.Researchers at UC Berkeley have identified over 150 such sites. In proof-of-concept experiments, a PDZ protein interaction domain has been intercalated and increased functionality without decreasing Cas9 nuclease activity. In further experiments, the internal insertion sites have been used to alter Cas9 activity in an allosteric manner, effectively creating tunable Cas9.

Catalytic Synthesis Of Fluorinated Anilines

Molecules containing aniline and aniline derivatives are common in the pharmaceutical, agrochemical, and pigment industries and numerous methods for the preparation of anilines have been reported.  Aniline derivatives containing electron-withdrawing substituents are more valuable in medicinal chemistry because anilines are prone to oxidation.  The past methods to obtain fluorinated anilines, which also mitigate oxidation, have been limited and the yields were moderate.   UC Berkeley researchers have developed a reaction for the coupling of primary fluoroalkylamines with aryl bromides and aryl chlorides and occur in the presence of functional groups that are typically not tolerated by C-N coupling reactions. The reaction yield is high and can be conducted with low catalyst loadings for most substrates.  

Methods For High Signal-To-Noise Imaging Of Chromosomal Loci In Cells Using Fluorescent Cas9

Cas9 is an endonuclease that binds complementary target DNA and generates site-specific breaks using two conserved nuclease domains. By inactivating both nuclease domains, dCas9 is produced, which functions as a programmable DNA binding protein. Current methods use dCas9-GFP fusions to image chromosomal loci, but have insufficient signal-to-noise ratio and often misidentify loci. UC Berkeley researchers have engineered a Cas9 variant that can be labeled with small molecule fluorescent dyes. This variant utilizes a conformational change in Cas9 to provide highly specific identification of chromosomal loci, and has been shown to work in a proof-of-principle experiment using Förster resonance energy transfer (FRET) pairs.

Live Imaging of Corneal Lymphatic Vessels

Lymphatic research is an explosive field of new discovery in recent years. Lymphatic dysfunction has been found in a wide array of disorders which include but are not limited to cancers and tumors, inflammation, infection, autoimmune diseases, dry eye, chemical burn, and tissue or organ transplant rejection, etc. The cornea provides an optimal site for lymphatic research due to its accessible location, transparent nature, and lymphatic-free but inducible features. Because there are no pre-existing vessels to consider in this unique tissue, it is exceptionally straightforward and accurate to assess lymphatic events (from formation to maturation and regression) in the cornea. Since lymphatic vessels are not easily visible as blood vessels, previous studies using the cornea have relied on traditional immunohistochemistry assays with dead tissues. Currently, there are no means of direct and harmless visualization of lymphatic vessels within live cornea. Investigators at University of California at Berkeley have addressed this challenge by developing the first live imaging of corneal lymphatic vessels. Lymphatic specific dye is injected into the subconjunctival space to visualize lymphatic vessels at various stages in the cornea under a fluorescence stereo, confocal, or two-photon microscope. Moreover, lymphatic vessels can be visualized in different colors to produce two, three, and four-dimensional images or live videos at a molecular level. The investigators have demonstrated a proof of principle in live mouse cornea. The technique allows time course tracking of dynamic lymphatic processes within the same tissue or subject over a short or long period of time, and can be ideally used to assess the progression of disease development and the effect of drug treatment. Live imaging of corneal lymphatic vessels allows visualization of lymphatic vessels in their natural morphology, state, and interactions with the local environment. This noninvasive method of live imaging of corneal lymphatic vessels is readily applicable to patient examination and the lymphatic dye of dextran is bio-degradable and harmless to human health.


Plant growth and development depends on the coordination of gene expression in a tissue-, temporal-, or signal-dependent manner. Often, the complex expression pattern observed for a given gene derives from regulation at both the transcriptional and post-transcriptional levels. This multi-layered approach to gene control is part of what makes plants robust, adaptable, and efficient as living photosynthetic systems. Complex gene regulation requires a multi-level approach. Current technology for transgene regulation in plants is based almost exclusively on transcriptional activation. For example, tissue-specific and stress-responsive promoters are extensively employed for inducible gene regulation, though many of these conditional promoters are species-specific. Alternative splicing of mRNA is an important mechanism of gene regulation in plants. The two possible consequences of alternative splicing of an mRNA are: (i) to change the protein coding sequence, by inserting or deleting sequences, or by shifting the reading frame, or (ii) to change the fate of the mRNA, by inserting or deleting sequences that target the RNA for degradation, localization, or other processes. This research has identified how to use RNA elements that regulate alternative splicing of mRNAs for inducible control of gene expression.

Maize plants with reduced gene silencing

Normal 0 0 1 95 543 UC Berkeley 4 1 666 11.1282 0 0 0 This invention relates to genes and methods to control eukaryotic mechanisms responsible for both the genesis and maintenance of heritable phenotypic variation. Paramutations represent specific types of epigenetic alterations that can be stably inherited without altering the DNA sequence. Several genes are known to be involved in regulating the paramutant state in maize plants. It is currently thought that these allelic interactions cause structural alterations to the chromatin. This invention relates to the use of the rmr1 gene for reducing or mitigating gene silencing in a transgenic maize plant and also to reduce or mitigate inbreeding depression

Dna Recombination In Eukaryotic Cells By The Bacteriophage Phic31 Recombination System

This invention provides methods for obtaining specific and stable integration of nucleic acids into eukaryotic cells. The method is an alternative to the widely used CRE-LOX method. The invention makes use of site-specific recombination systems that use prokaryotic recombinase polypeptides, such as the phiC31 recombinase, that can mediate recombination between the recombination sites, but not between hybrid recombination sites that are formed upon the recombination. Thus, the recombination is irreversible in the absence of additional factors. Eukaryotic cells that contain the recombinase polypeptides, or genes that encode the recombinases, are also provided. See also: MGG Molecular Genetics and Genomics. August, 2001. 265(6):1031-1038. Thomason, L. C.; Calendar, R.; Ow, D. W. Gene insertion and replacement in Schizosaccharomyces pombe mediated by the Streptomyces bacteriophagevariant phiC31 site-specific recombination system. Abstract: The site-specific recombination system used by the Streptomyces bacteriophage variant phiC31 was tested in the fission yeast Schizosaccharomyces pombe. A target strain with the phage attachment site attP inserted at the leu1 locus was co-transformed with one plasmid containing the bacterial attachment site attB linked to a ura4+ marker, and a second plasmid expressing the variant phiC31 integrase gene. High-efficiency transformation to the Ura+ phenotype occurred when the integrase gene was expressed. Southern analysis revealed that the attB-ura4+ plasmid integrated into the chromosomal attP site. Sequence analysis showed that the attBXattP recombination was precise. In another approach, DNA with a ura4+ marker flanked by two attB sites in direct orientation was used to transform S. pombe cells bearing an attP duplication. The variant phiC31 integrase catalyzed two reciprocal cross-overs, resulting in a precise gene replacement. The site-specific insertions are stable, as no excision (the reverse reaction) was observed on maintenance of the integrase gene in the integrant lines. The irreversibility of the variant phiC31 site-specific recombination system sets it apart from other systems currently used in eukaryotic cells, which reverse readily. Deployment of the variant phiC31 recombination provides new opportunities for directing transgene Y and chromosome rearrangements in eukaryotic systems.

Photosynthetic Hydrogen Production Using Algae

Hydrogen gas is considered to be the ideal fuel for combating environmental degradation. However, the biggest obstacle to hydrogen replacing petroleum as the world's primary source of energy is the high cost of cleanly producing this gas. The most cost-effective current method for producing H2 is to use nuclear energy -- but that has environmental issues. Likewise, using solar power is not cost-effective and using wind power is limited to a few regions. To address this challenge, researchers at the University of California, Berkeley have developed a photosynthetic method for producing H2. This patented H2 production method is based on depriving algae of sulfur which in turn inhibits oxygen flow and augments its natural H2 production. Using a bioreactor comprised of a network of sealed tubes for cultivating algae and extracting pure H2, researchers were able to produce the gas for about $0.31 per kilowatt-hour. That is much higher than natural gas-fired methods that produce H2 for about $0.05 per kilowatt-hour. However, the Berkeley team is pursuing research to address bottlenecks in this photosynthetic process which would in turn improve efficiency and reduce costs. These cost savings from the more efficient photosynthetic process along with refinements to the bioreactor design could make this algae production method cost competitive with the natural gas-fired production approach.

Cereal Transformation Of Using Cultured Organogenic Tissue

A novel transformation method for transforming many commercial genotypes of cereals has been developed. The method incolves using meristematic tissues which include vegetative shoot meristems and young leaf bases as initial explants and the induction of a direct organogenesis pathway for in vitro proliferation and plant regeneration. Use of this pathway results in less impact and destabilizaton of the methylation state of the genomic DNa during invitro growth and reduces the impact of somaclonal variation and instability of transgene expression. References; S. Zhang, et al. 1999. Genetic transformation of commercial cultivars of oat (Avena sativa L.) and barley (Hordeum vulgare L.) using in vitro shoot meristem cultures derived from germinated seedlings. Plant Cell Reports. 18:959-66. S. Zhang, et al., 1998. Expression of CDC2Zm an dknotted 1 during in=vitro axillary shoot meristem proliferation and adventitious shoot meristem formation in maize (Zea mays L.) and barley (Hordeum vulgare L.). Planta. 204:542-9

Methods For The Transformation Of Monocots Using Green Embryogenic Callus

Investigators at the University of California, Berkeley have developed improved compositions and methods for the transformation of monocots from embryogenic callus. Such methods include, for example, use of an intermediate incubation medium after callus induction to increase the competence of the transformed cells for regeneration; dim light conditions during early phases of selection; use of green callus tissue as a target for microprojectile bombardment; and media with optimized levels of phytohormones and copper concentrations.

Monoclonal Antibodies For Identification Of Prunus Necrotic Ringspot Virus And Related Stone Fruit Viruses

Prunus Necrotic Ringspot virus (PNRV) and its variants infect and have pathologic effects on a wide variety of stone fruit trees, apples, hops, and roses. This large economic impact necessitates a simple, sensitive, and highly reliable diagnostic test for these viruses in leaves, active and dormant buds, and other tissues. UC Berkeley researchers have developed 405 hybridoma lines specific for PNRV. To date, five of these have been investigated in depth. In initial tests by four laboratories, these antibodies reacted with a variety of PNRV isolates from California that commercially available antisera and monoclonal antibodies recognized more weakly or not at all.

Determination of Bioproduct Content in Live Cell Cultures for Industrial Applications

There is great variability among different organisms in their ability to naturally or artificially synthesize and accumulate lipids, hydrocarbons, and polymers. Consequently, many organisms must be screened in order to achieve the desired maximal bio-product accumulation. After an ideal organism is selected, its product content can vary with lifecycle stage, cultivation conditions, cellular stress and/or time. This variability must be understood and controlled during R&D, process development and manufacturing scale-up in order to maximize product yields. The above process of screening and development can be time-consuming and consequently costly.  To address this situation, scientists at UC Berkeley have developed a method for quick and precise estimation of lipid, hydrocarbon or biopolymer content in live cells -- whether grown as single cells or in colonies. This method can be used for screening a variety of microorganisms for product accumulation (microorganism prospecting), and to check yields throughout the production process -- allowing for more rapid improvement of production methods and shortened R&D timelines.

Wireless Systems For Process Monitoring

Chemical, biochemical and agricultural processes such as fermentation, vaccine production, require close monitoring for quality control and process optimization. For some processes, production of gaseous emissions must be constantly monitored to insure worker safety or compliance with environmental regulations. Systems for many of these process monitoring applications can be very expensive and inflexible; for example where deployment requires fixed wiring for power supplies and data transmission. Systems can also be difficult to retrofit when existing facilities are used for new processing operations, or sensors must be added for monitoring new or different gaseous species. Researchers at the University of California, Berkeley have developed a wireless monitoring system for liquid processing operations. The system is designed to monitor a variety of processes, including the fermentation of wine, beer, and spirits. The system allows for rapid deployment of self-organizing sensor networks for the monitoring within production equipment (such as fermentation tanks or vats) as well as at other locations within and outside the production facility. The network can also be expanded to monitor post processing steps such as bottling or packaging. The network utilizes small, wireless sensors that are low cost and highly scalable, and the system allow for rapidly deployment into evolving liquid processing environments.

A Mutant Of The Green Alga Dunaliella Salina Accumulates Zeaxanthin, A High-value Bio-product

A novel mutant of of the halotolerant unicellular green alga Dunaliella salinia lacks a number of the beta-branch xanthophylls but accumulates zeaxanthin. Biochemical analysis suggests that zeaxanthin substitutes for the beta-branched xanthophylls in the mutant strain. This mutant may provide a biological source for the production of zeaxanthin, a high value bio-product. Ref.: E. Jin, & et al. 2003. Biotechnol Bioeng 81:115-124

A Universal, Light-switchable Gene Promoter System

Synopsis: This invention consists of an artificial promoter system that can be fused upstream of any desired gene, enabling reversible and light-switchable induction or repression of gene expression in any suitable host cell. New data to be filed in a provisional patent application demonstrates optimized expression conditions and a "switching off" mechanism in addition to the "switching on" mechanism.

Genetic Functions Required For Gene Silencing In Maize

Hollick, Jay B.; Chandler, Vicki L. Genetic factors required to maintain repression of a paramutagenic maize pl1 allele. Genetics. January, 2001. 157(1):369-378. Abstract: A genetic screen identified two novel gene functions required to maintain mitotically and meiotically heritable gene silencing associated with paramutation of the maize purple plant 1 (pl1) locus. Paramutation at pl1 leads to heritable alterations of pl1 gene regulation; the Pl-Rhoades (Pl-Rh) allele, which typically confers strong pigmentation to juvenile and adult plant structures, changes to a lower expression state termed Pl'-mahogany (Pl'). Paramutation spontaneously occurs at low frequencies in Pl-Rh homozygotes but always occurs when Pl-Rh is heterozygous with Pl'. We identified four mutations that caused increased Pl' pigment levels. Allelism tests revealed that three mutations identified two new maize loci, required to maintain repression 1 (rmr1) and rmr2 and that the other mutation represents a new allele of the previously described mediator of paramutation 1 (mop1) locus. RNA levels from Pl' are elevated in rmr mutants and genetic tests demonstrate that Pl' can heritably change back to Pl-Rh in rmr mutant individuals at variable frequencies. Pigment levels controlled by two pl1 alleles that do not participate in paramutation are unaffected rmr mutants. These results suggest that RMR functions are intimately involved in maintaining the repressed expression state of paramutant Pl' alleles. Despite strong effects on Pl' repression, rmr mutant plants have no gross developmental abnormalities even after several generations of inbreeding, implying that RMR1 and RMR2 functions are not generally required for developmental homeostasis. also see: Dorweiler, Jane E.; Carey, Charles C.; Kubo, Kenneth M.; Hollick, Jay B.; Kermicle, Jerry L.; Chandler, Vicki L. mediator of paramutation1 is required for establishment and maintenance of paramutation at multiple maize loci. Plant Cell. November, 2000. 12(11):2101-2118. Abstract: Paramutation is the directed, heritable alteration of the expression of one allele when heterozygous with another allele. Here, the isolation and characterization of a mutation affecting paramutation, mediator of paramutation1-1 (mop1-1), are described. Experiments demonstrate that the wild-type gene Mop1 is required for establishment and maintenance of the paramutant state. The mop1-1 mutation affects paramutation at the multiple loci tested but has no effect on alleles that do not participate in paramutation. The mutation does not alter the amounts of actin and ubiquitin transcripts, which suggests that the mop1 gene does not encode a global repressor. Maize plants homozygous for mop1-1 can have pleiotropic developmental defects, suggesting that mop1-1 may affect more genes than just the known paramutant ones. The mop1-1 mutation does not alter the extent of DNA methylation in rDNA and centromeric repeats. The observation that mop1 affects paramutation at multiple loci, despite major differences between these loci in their gene structure, correlations with DNA methylation, and stability of the paramutant state, suggests that a common mechanism underlies paramutation. A protein-based epigenetic model for paramutation is discussed.

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