The Experts below are selected from a list of 324 Experts worldwide ranked by ideXlab platform
Wolfgang Spielmeyer - One of the best experts on this subject based on the ideXlab platform.
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an accurate DNA Marker assay for stem rust resistance gene sr2 in wheat
Theoretical and Applied Genetics, 2011Co-Authors: Rohit Mago, Gina Brownguedira, Susanne Dreisigacker, James Breen, Ravi P Singh, R Appels, E S Lagudah, J G Ellis, Wolfgang SpielmeyerAbstract:The stem rust resistance gene Sr2 has provided broad-spectrum protection against stem rust (Puccinia graminis Pers. f. sp. tritici) since its wide spread deployment in wheat from the 1940s. Because Sr2 confers partial resistance which is difficult to select under field conditions, a DNA Marker is desirable that accurately predicts Sr2 in diverse wheat germplasm. Using DNA sequence derived from the vicinity of the Sr2 locus, we developed a cleaved amplified polymorphic sequence (CAPS) Marker that is associated with the presence or absence of the gene in 115 of 122 (95%) diverse wheat lines. The Marker genotype predicted the absence of the gene in 100% of lines which were considered to lack Sr2. Discrepancies were observed in lines that were predicted to carry Sr2 but failed to show the CAPS Marker. Given the high level of accuracy observed, the Marker provides breeders with a selection tool for one of the most important disease resistance genes of wheat.
Karin R Deal - One of the best experts on this subject based on the ideXlab platform.
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molecular characterization of a diagnostic DNA Marker for domesticated tetraploid wheat provides evidence for gene flow from wild tetraploid wheat to hexaploid wheat
Molecular Biology and Evolution, 2006Co-Authors: Jan Dvorak, Eduard Akhunov, Alina R Akhunov, Karin R DealAbstract:All forms of domesticated tetraploid wheat (Triticum turgidum, genomes AABB) are nearly monomorphic for restriction fragment length polymorphism (RFLP) haplotype a at the Xpsr920 locus on chromosome 4A (Xpsr920-Ala), and wild tetraploid wheat is monomorphic for haplotype b. The Xpsr920-Ala/b dimorphism provides a molecular Marker for domesticated and wild tetraploid wheat, respectively. Hexaploid wheat (Triticum aestivum, genomes AABBDD) is polymorphic for the 2 haplotypes. Bacterial artificial chromosome (BAC) clones hybridizing with PSR920 were isolated from Triticum urartu (genomes AA), Triticum monococcum (genomes A m A m ), and T. turgidum ssp. durum (genomes AABB) and sequenced. PSR920 is a fragment of a putative ATP binding cassette (ABC) transporter gene (designated ABCT-1). The wheat ABCT-1 gene is more similar to the T. urartu gene than to the T. monococcum gene and diverged from the T. urartu gene about 0.7 MYA. The comparison of the sequence of the wheat A genome BAC clone with that of the T. urartu BAC clone provides the first insight into the microsynteny of the wheat A genome with that of T. urartu. Within 103 kb of orthologous intergenic space, 37 kb of new DNA has been inserted and 36 kb deleted leaving 49.7% of the region syntenic between the clones. The nucleotide substitution rate in the syntenic intergenic space has been 1.6 X 10 -8 nt -1 year -1 , which is, respectively, 4 and 3 times as great as nucleotide substitution rates in the introns and the third codon positions of the juxtaposed gene. The RFLP is caused by a miniature inverted transposable element (MITE) insertion into intron 18 of the ABCT-A1 gene. Polymerase chain reaction primers were developed for the amplification of the MITE insertion site and its sequencing. The T. aestivum ABCT-A1a haplotype is identical to the haplotype of domesticated tetraploid wheat, and the ABCT-A1 b haplotype is identical to that of wild tetraploid wheat. This finding shows for the first time that wild tetraploid wheat participated in the evolution of hexaploid wheat. A dine of the 2 haplotype frequencies exists across Euro-Asia in T. aestivum. It is suggested that T. aestivum in eastern Asia conserved the gene pool of the original T. aestivum more than wheat elsewhere.
Steven S Xu - One of the best experts on this subject based on the ideXlab platform.
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targeted introgression of a wheat stem rust resistance gene by DNA Marker assisted chromosome engineering
Genetics, 2011Co-Authors: Daryl L Klindworth, Timothy L Friesen, Shiaoman Chao, Steven S XuAbstract:Chromosome engineering is a useful strategy for transfer of alien genes from wild relatives into modern crops. However, this strategy has not been extensively used for alien gene introgression in most crops due to low efficiency of conventional cytogenetic techniques. Here, we report an improved scheme of chromosome engineering for efficient elimination of a large amount of goatgrass (Aegilops speltoides) chromatin surrounding Sr39, a gene that provides resistance to multiple stem rust races, including Ug99 (TTKSK) in wheat. The wheat ph1b mutation, which promotes meiotic pairing between homoeologous chromosomes, was employed to induce recombination between wheat chromosome 2B and goatgrass 2S chromatin using a backcross scheme favorable for inducing and detecting the homoeologous recombinants with small goatgrass chromosome segments. Forty recombinants with Sr39 with reduced surrounding goatgrass chromatin were quickly identified from 1048 backcross progenies through disease screening and molecular Marker analysis. Four of the recombinants carrying Sr39 with a minimal amount of goatgrass chromatin (2.87–9.15% of the translocated chromosomes) were verified using genomic in situ hybridization. Approximately 97% of the goatgrass chromatin was eliminated in one of the recombinants, in which a tiny goatgrass chromosome segment containing Sr39 was retained in the wheat genome. Localization of the goatgrass chromatin in the recombinants led to rapid development of three molecular Markers tightly linked to Sr39. The new wheat lines and Markers provide useful resources for the ongoing global effort to combat Ug99. This study has demonstrated great potential of chromosome engineering in genome manipulation for plant improvement.
Rohit Mago - One of the best experts on this subject based on the ideXlab platform.
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an accurate DNA Marker assay for stem rust resistance gene sr2 in wheat
Theoretical and Applied Genetics, 2011Co-Authors: Rohit Mago, Gina Brownguedira, Susanne Dreisigacker, James Breen, Ravi P Singh, R Appels, E S Lagudah, J G Ellis, Wolfgang SpielmeyerAbstract:The stem rust resistance gene Sr2 has provided broad-spectrum protection against stem rust (Puccinia graminis Pers. f. sp. tritici) since its wide spread deployment in wheat from the 1940s. Because Sr2 confers partial resistance which is difficult to select under field conditions, a DNA Marker is desirable that accurately predicts Sr2 in diverse wheat germplasm. Using DNA sequence derived from the vicinity of the Sr2 locus, we developed a cleaved amplified polymorphic sequence (CAPS) Marker that is associated with the presence or absence of the gene in 115 of 122 (95%) diverse wheat lines. The Marker genotype predicted the absence of the gene in 100% of lines which were considered to lack Sr2. Discrepancies were observed in lines that were predicted to carry Sr2 but failed to show the CAPS Marker. Given the high level of accuracy observed, the Marker provides breeders with a selection tool for one of the most important disease resistance genes of wheat.
Charlotte M Niemeyer - One of the best experts on this subject based on the ideXlab platform.
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Immuno-PCR with digital readout
Biochemical and Biophysical Research Communications, 2017Co-Authors: Hendrik Schröder, Mark Spengler, Maximilian Grösche, Michael Adler, Charlotte M NiemeyerAbstract:Immuno-PCR (IPCR) combines the versatile ELISA antigen detection with ultrasensitive PCR signal amplification, thereby enabling the highly sensitive detection of a broad range of targets with a typically very large dynamic detection range. The quantification of the antigen is usually achieved by real-time PCR, which provides a correlation between the target concentration and amplified DNA Marker. We here report on the implementation of digital droplet PCR as a means for direct quantification of DNA copies to enable the highly sensitive detection of protein bioMarkers. To this end, two alternative approaches, based on either magnetic microbead-based IPCR or a microplate-release IPCR were tested. The latter format worked well and revealed an extraordinary high robustness and sensitivity. While rtIPCR already fulfills typical immunoassay acceptance criteria, ddIPCR enables improved accuracy and precision of the assay because signal response and analyte concentrations are directly correlated. The utility of the novel ddIPCR technology is demonstrated at the example of two cytokines, interleukin 2 and interleukin 6 (IL2, IL6, respectively), with an overall average CV% of 5.0 (IL2) and 7.4 (IL6).
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Highly sensitive ligand-binding assays in pre-clinical and clinical applications: Immuno-PCR and other emerging techniques
Analyst, 2015Co-Authors: Mark Spengler, Michael Adler, Charlotte M NiemeyerAbstract:Emerging state-of-the-art ligand-binding assay technologies for pharmaceutical sample testing are surveyed, which reveal enhanced analytical sensitivity over classical ELISA formats.Recombinant DNA technology and corresponding innovations in molecular biology, chemistry and medicine have led to novel therapeutic biomacromolecules as lead candidates in the pharmaceutical drug development pipelines. While monoclonal antibodies and other proteins provide therapeutic potential beyond the possibilities of small molecule drugs, the concomitant demand for supportive bioanalytical sample testing creates multiple novel challenges. For example, intact macromolecules can usually not be quantified by mass-spectrometry without enzymatic digestion and isotopically labeled internal standards are costly and/or difficult to prepare. Classical ELISA-type immunoassays, on the other hand, often lack the sensitivity required to obtain pharmacokinetics of low dosed drugs or pharmacodynamics of suitable bioMarkers. Here we summarize emerging state-of-the-art ligand-binding assay technologies for pharmaceutical sample testing, which reveal enhanced analytical sensitivity over classical ELISA formats. We focus on immuno-PCR, which combines antibody specificity with the extremely sensitive detection of a tethered DNA Marker by quantitative PCR, and alternative nucleic acid–based technologies as well as methods based on electrochemiluminescence or single-molecule counting. Using case studies, we discuss advantages and drawbacks of these methods for preclinical and clinical sample testing.