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Research Interests: Biology and Food Allergy
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ABSTRACT Successful efforts describing in vitro culturing, regeneration, and transformation of grain sorghum were reported, using particle bombardment, as early as 1993, and with Agrobacterium tumefaciens in 2000. Reported transformation... more
ABSTRACT Successful efforts describing in vitro culturing, regeneration, and transformation of grain sorghum were reported, using particle bombardment, as early as 1993, and with Agrobacterium tumefaciens in 2000. Reported transformation efficiencies via Agrobacterium routinely range from 1 to 2%. Recently, such efficiencies via Agrobacterium in several plant species were improved with the use of heat and centrifugation treatments of explants prior to infection. Here, we describe the successful use of heat pretreatment of immature embryos (IEs) prior to Agrobacterium inoculation to increase routine transformation frequencies of a single genotype, P898012, to greater than 7%. This reproducible frequency was calculated as numbers of independently transformed IEs, confirmed by PCR, western, and DNA hybridization analysis, that produced fertile transgenic plants, divided by total numbers of infected IEs.
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Characterization of mutant alleles provides valuable insights into the genetics of normal physiological functions. Complementing the study of mutations generated randomly with chemicals or irradiation, technical advances in the field of... more
Characterization of mutant alleles provides valuable insights into the genetics of normal physiological functions. Complementing the study of mutations generated randomly with chemicals or irradiation, technical advances in the field of genomics and the explosion of available gene sequences provide tools that lead to increasingly detailed characterizations of individual genes and their physiological roles.
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Sugar beet (Beta vulgaris L.) is an important industrial crop, being one of only two plant sources from which sucrose (ie, sugar) can be economically produced. Despite its relatively short period of cultivation (ca. 200 years), its yield... more
Sugar beet (Beta vulgaris L.) is an important industrial crop, being one of only two plant sources from which sucrose (ie, sugar) can be economically produced. Despite its relatively short period of cultivation (ca. 200 years), its yield and quality parameters have been significantly ...
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Homozygous lines of barley overexpressing a wheat thioredoxin h transgene (up to 30-fold) were generated earlier by using a B 1 -hordein promoter with a signal peptide sequence for targeting to the protein body and found to be enriched in... more
Homozygous lines of barley overexpressing a wheat thioredoxin h transgene (up to 30-fold) were generated earlier by using a B 1 -hordein promoter with a signal peptide sequence for targeting to the protein body and found to be enriched in starch debranching enzyme (pullulanase). Here, we describe the effect of biochemically active, overexpressed thioredoxin h on germination and the onset of α-amylase activity. Relative to null segregant controls lacking the transgene, homozygotes overexpressing thioredoxin h effected ( i ) an acceleration in the rate of germination and appearance of α-amylase activity with a 1.6- to 2.8-fold increase in gibberellin A 1 (GA 1 ) content; ( ii ) a similar acceleration in the appearance of the α-amylase activity in deembryonated transgenic grain incubated with gibberellic acid; ( iii ) a 35% increase in the ratio of relative reduction (abundance of SH) of the propanol soluble proteins (hordein I fraction); and ( iv ) an increase in extractable and solub...
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The homeobox gene, knotted1, (kn1) is expressed in shoot meristems and is required for maintaining indeterminacy and preventing cellular differentiation. Awns, extensions of the bract-like lemma found in all grass inflorescences, are... more
The homeobox gene, knotted1, (kn1) is expressed in shoot meristems and is required for maintaining indeterminacy and preventing cellular differentiation. Awns, extensions of the bract-like lemma found in all grass inflorescences, are normally determinate structures. We show that ectopic expression of kn1 in the barley awn is sufficient to direct the development of ectopic meristems, forming inflorescence-like structures. This homeotic transformation is similar to the phenotype produced by misexpression of the barley hvknox3 gene, associated with the dominant Hooded mutant (Müller, K. J., Romano, N., Gerstner, O., Garcia-Maroto, F., Pozzi, C., Salamini, F. and Rohde, W. (1995) Nature 374, 727-730). We suggest that the inverse polarity of the ectopic flowers seen in Hooded and transgenic kn1 plants results from the transformation of the awn into reiterative inflorescence axes. We observed that the protein and mRNA localization of the transgene, driven by a constitutive promoter, is si...
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... with respect to the transformation of maize elite inbreds over those methods previously described in which direct bombardment of the SAM in coleoptilar ... Lowe K, Ross M, Sandahl G, Miller M, Hoerster G, Church L, Tagliani L, Bond D,... more
... with respect to the transformation of maize elite inbreds over those methods previously described in which direct bombardment of the SAM in coleoptilar ... Lowe K, Ross M, Sandahl G, Miller M, Hoerster G, Church L, Tagliani L, Bond D, Gordon-Kamm WJ (1997) Transformation of ...
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Photosynthesis Research 17:23-56 (1988) © Kluwer Academic Publishers, Dordrecht - Printed in the Netherlands ... Characterization of phycobiliprotein and linker polypeptide genes in Fremyella diplosiphon and their regulated expression... more
Photosynthesis Research 17:23-56 (1988) © Kluwer Academic Publishers, Dordrecht - Printed in the Netherlands ... Characterization of phycobiliprotein and linker polypeptide genes in Fremyella diplosiphon and their regulated expression during complementary chromatic adaptation*
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Genetic transformation using shoot meristematic cultures (SMCs) derived from germinated seedlings is established in commercial varieties of oat cv 'Garry' and barley cv 'Harrington'. Six-month-old SMCs of oat were... more
Genetic transformation using shoot meristematic cultures (SMCs) derived from germinated seedlings is established in commercial varieties of oat cv 'Garry' and barley cv 'Harrington'. Six-month-old SMCs of oat were induced on MPM and bombarded with bar and uidA; 9-month-old SMCs of barley were induced on an improved medium (MPM-MC) containing maltose and high levels of copper and bombarded with bar/nptII and uidA. After 3–4 months on selection, seven independent transgenic lines of oat were obtained, two lines of barley. All transgenic lines produced T0 plants; five lines of oat and one line of barley were self-fertile, and the other barley line produced T1 seed when out-crossed. Both Mendelian and non-Mendelian segregation ratios of transgene expression were observed in T1 and T2 progeny of transgenic oat. Normal as well as low physical transmission of the transgenes was also seen in T1 and T2 progeny of oat. The bar-containing line of barley showed stable transg...
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The expression level of bar, which encodes phosphinothricin acetyltransferase (PAT), was correlated with the inviability of barley hybrids between 20 Golden Promise-derived transgenic lines (Ds-bar lines) and a specialized genetic marker... more
The expression level of bar, which encodes phosphinothricin acetyltransferase (PAT), was correlated with the inviability of barley hybrids between 20 Golden Promise-derived transgenic lines (Ds-bar lines) and a specialized genetic marker stock, Oregon Wolfe Barley Dominant ( ...
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Barley is the cornerstone of the malting and brewing industry. It is known that 250 quantitative trait loci (QTLs) of the grain are associated with 19 malting-quality phenotypes. However, only a few of the contributing genetic components... more
Barley is the cornerstone of the malting and brewing industry. It is known that 250 quantitative trait loci (QTLs) of the grain are associated with 19 malting-quality phenotypes. However, only a few of the contributing genetic components have been identified. One of these, on chromosome 4H, contains a major malting QTL, QTL2, located near the telomeric region that accounts, respectively, for 28.9% and 37.6% of the variation in the β-glucan and extract fractions of malt. In the current study, we dissected the QTL2 region using an expression- and microsynteny-based approach. From a set of 22 expressed sequence tags expressed in seeds at the malting stage, we identified a candidate gene, TLP8 (thaumatin-like protein 8), which was differentially expressed and influenced malting quality. Transcript abundance and protein profiles of TLP8 were studied in different malt and feed varieties using quantitative PCR, immunoblotting, and enzyme-linked immunosorbent assay (ELISA). The experiments ...
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Plant transformation has enabled fundamental insights into plant biology and revolutionized commercial agriculture. Unfortunately, for most crops, transformation and regeneration remain arduous even after more than thirty years of... more
Plant transformation has enabled fundamental insights into plant biology and revolutionized commercial agriculture. Unfortunately, for most crops, transformation and regeneration remain arduous even after more than thirty years of technological advances. Genome editing provides new opportunities to enhance crop productivity, but relies on genetic transformation and plant regeneration, which are bottlenecks in the process. Herein we review the state of plant transformation and point to innovations needed to enable genome editing in crops. Plant tissue culture methods need optimization and simplification for efficiency and minimize time in culture. Currently, specialized facilities exist for crop transformation. Single cell and robotic techniques should be developed for high throughput genomic screens. Utilization of plant genes involved in developmental reprogramming, wound response, and/or homologous recombination could boost recovery of transformed plants. Engineering universal Agr...
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A number of parameters related to Agrobacte-rium-mediated infection were tested to optimize transformation frequencies of sorghum (Sorghum bicolor L.). A plasmid with a selectable marker, phosphomannose isomerase, and an sgfp reporter... more
A number of parameters related to Agrobacte-rium-mediated infection were tested to optimize transformation frequencies of sorghum (Sorghum bicolor L.). A plasmid with a selectable marker, phosphomannose isomerase, and an sgfp reporter gene was used. First, storing ...
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Thioredoxins (Trxs) are small ubiquitous regulatory disulfide proteins. Plants have an unusually complex complement of Trxs composed of six well-defined types (Trxs f, m, x, y, h, and o) that reside in different cell compartments and... more
Thioredoxins (Trxs) are small ubiquitous regulatory disulfide proteins. Plants have an unusually complex complement of Trxs composed of six well-defined types (Trxs f, m, x, y, h, and o) that reside in different cell compartments and function in an array of processes. ...
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The ecology of fungi lags behind that of plants and animals because most fungi are microscopic and hidden in their substrates. Here, we address the basic ecological process of fungal succession in nature using the microscopic, arbuscular... more
The ecology of fungi lags behind that of plants and animals because most fungi are microscopic and hidden in their substrates. Here, we address the basic ecological process of fungal succession in nature using the microscopic, arbuscular mycorrhizal fungi (AMF) that form essential mutualisms with 70-90% of plants. We find a signal for temporal change in AMF community similarity that is 40-fold stronger than seen in the most recent studies, likely due to weekly samplings of roots, rhizosphere and soil throughout the 17 weeks from seedling to fruit maturity and the use of the fungal DNA barcode to recognize species in a simple, agricultural environment. We demonstrate the patterns of nestedness and turnover and the microbial equivalents of the processes of immigration and extinction, that is, appearance and disappearance. We also provide the first evidence that AMF species co-exist rather than simply co-occur by demonstrating negative, density-dependent population growth for multiple ...
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Drought stress is a major obstacle to crop productivity, and the severity and frequency of drought are expected to increase in the coming century. Certain root-associated bacteria have been shown to mitigate the negative effects of... more
Drought stress is a major obstacle to crop productivity, and the severity and frequency of drought are expected to increase in the coming century. Certain root-associated bacteria have been shown to mitigate the negative effects of drought stress on plant growth, and manipulation of the crop microbiome is an emerging strategy for overcoming drought stress in agricultural systems, yet the effect of drought on the development of the root microbiome is poorly understood. Through 16S rRNA amplicon and metatranscriptome sequencing, as well as root metabolomics, we demonstrate that drought delays the development of the early sorghum root microbiome and causes increased abundance and activity of monoderm bacteria, which lack an outer cell membrane and contain thick cell walls. Our data suggest that altered plant metabolism and increased activity of bacterial ATP-binding cassette (ABC) transporter genes are correlated with these shifts in community composition. Finally, inoculation experime...
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Research Interests: Plant Biology, Transcription Factors, Comparative Study, Gene expression, In Situ Hybridization, and 15 moreMaize, Somatic Embryogenesis, Embryonic Development, Genotype, Seeds, Molecular Marker, Gramineae, Embryos, PLANT PROTEINS, Planta, Meristem, Gene expression profiling, Embryo Development, Somatic Cells, and Expression pattern
A rapid, efficient, and reproducible system to generate large numbers of independently transformed, self-fertile, transgenic bar- ley (Hordeum vurgare 1.) plants is described. lmmature zygotic embryos, young callus, and microspore-derived... more
A rapid, efficient, and reproducible system to generate large numbers of independently transformed, self-fertile, transgenic bar- ley (Hordeum vurgare 1.) plants is described. lmmature zygotic embryos, young callus, and microspore-derived embryos were bombarded with a plasmid containing bar and uidA either alone or in combination with another plasmid containing a barley yellow dwarf virus coat protein (BYDVcp) gene. A total
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Improved compositions and methods for transformation and regeneration of plants from embryogenic callus are disclosed that include, for example: use of an intermediate-incubation medium after callus induction to increase the competence of... more
Improved compositions and methods for transformation and regeneration of plants from embryogenic callus are disclosed that 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..
74) Attorney) Agent) or Firm lvlorrison & Foerster LLP (56) References Cited (57) ABSTRACT
536/231; 435/320.1; 435/410; 435/252.1; 435/69.1; 435/183; 800/298 3/2003 Lanahan et al. FOREIGN PATENT DOCUMENTS JP 62-61783 9/1994 W0 WO 96/03505 2/1996 W0 WO 00/36126 6/2000 OTHER PUBLICATIONS Gilberger et al. al.... more
536/231; 435/320.1; 435/410; 435/252.1; 435/69.1; 435/183; 800/298 3/2003 Lanahan et al. FOREIGN PATENT DOCUMENTS JP 62-61783 9/1994 W0 WO 96/03505 2/1996 W0 WO 00/36126 6/2000 OTHER PUBLICATIONS Gilberger et al. al. "Characterization of Wheat thioredoXin h cDNA and production of an active Triticum aestivum protein in Escherichia coli " Eur. J. Biochem, 252: 314—324 (1998). Shi et al. Plant Molecular Biology 32: 653—662 (1996). Dai, Shaodong et al. (1996) "Crystal Structure of Arabia'op sis thaliana NADPH Dependent Thioredoxin Reductase at 2.5 A Resolution" J. Mol. Biol. 264: 1044—1057. Jacquot, Jean—Pierre et al. (1994) "Arabia'opsis thaliana NAPHP Thioredoxin Redutase" J. Mol. Biol. 235: 1357—1363. * cited by examiner Primary Examiner—Juliet C. SWitZer (74) Attorney, Agent, or Firm—Morrison & Foerster LLP (57) ABSTRACT The present invention provides barley thioredoXin h nucleic acids and NADP-thioredoxin reductase nucleic acids, the respective...
Transgenic monocot plants and seeds comprising monocot seed-specific promoters from seed-storage protein genes and monocot seed-specific targets sequences of expression of heterologous proteins in subcellular compartments to protect the... more
Transgenic monocot plants and seeds comprising monocot seed-specific promoters from seed-storage protein genes and monocot seed-specific targets sequences of expression of heterologous proteins in subcellular compartments to protect the protein are disclosed. The transgenic monocot plants include maize, wheat, rice and barley. Also disclosed are methods of producing transgenic monocot seed from the transgenic plants.
Barley (Hordeum vulgare L.) possesses a large and highly repetitive genome of 5.1 Gb that has hindered the development of a complete sequence. In 2012, the International Barley Sequencing Consortium released a resource integrating... more
Barley (Hordeum vulgare L.) possesses a large and highly repetitive genome of 5.1 Gb that has hindered the development of a complete sequence. In 2012, the International Barley Sequencing Consortium released a resource integrating whole-genome shotgun sequences with a physical and genetic framework. However, because only 6,278 BACs in the physical map were sequenced, fine structure was limited. To gain access to the gene-containing portion of the barley genome at high resolution, we identified and sequenced 15,622 BACs representing the minimal tiling path of 72,052 physical-mapped gene-bearing BACs. This generated ~1.7 Gb of genomic sequence containing an estimated 2/3 of all Morex barley genes. Exploration of these sequenced BACs revealed that although distal ends of chromosomes contain most of the gene-enriched BACs and are characterized by high recombination rates, there are also gene-dense regions with suppressed recombination. We made use of published map-anchored sequence data...