摘要:
Disclosed are methods and materials for assembling long polynucleotides from synthetic oligonucleotides. The use of synthetic oligonucleotides permits non-natural design of sequences. The oligonucleotides used for construction may be relatively short, according to practicalities of nucleotide synthesis. They are assembled using a ligase which is operative over a range of temperatures, i.e., is thermostable. The method and oligonucleotides are designed such that the melting temperature of the strands to be hybridized is set at a number of selected specific temperatures for each group of oligonucleotides to be hybridized and ligated. Hybridization and ligation take place at or near the melting temperature, so that each succeeding ligation is governed by a temperature that will prevent hybridization if any mismatches are present.
摘要:
High throughput screening of crystallization of a target material is accomplished by simultaneously introducing a solution of the target material into a plurality of chambers of a microfabricated fluidic device. The microfabricated fluidic device is then manipulated to vary the solution condition in the chambers, thereby simultaneously providing a large number of crystallization environments. Control over changed solution conditions may result from a variety of techniques, including but not limited to metering volumes of crystallizing agent into the chamber by volume exclusion, by entrapment of volumes of crystallizing agent determined by the dimensions of the microfabricated structure, or by cross-channel injection of sample and crystallizing agent into an array of junctions defined by intersecting orthogonal flow channels.
摘要:
The present invention relates to microfluidic devices and methods facilitating the growth and analysis of crystallized materials such as proteins. In accordance with one embodiment, a crystal growth architecture is separated by a permeable membrane from an adjacent well having a much larger volume. The well may be configured to contain a fluid having an identity and concentration similar to the solvent and crystallizing agent employed in crystal growth, with diffusion across the membrane stabilizing that process. Alternatively, the well may be configured to contain a fluid having an identity calculated to affect the crystallization process. In accordance with the still other embodiment, the well may be configured to contain a material such as a cryo-protectant, which is useful in protecting the crystalline material once formed.
摘要:
High throughput screening of crystallization of a target material is accomplished by simultaneously introducing a solution of the target material into a plurality of chambers of a microfabricated fluidic device. The microfabricated fluidic device is then manipulated to vary the solution condition in the chambers, thereby simultaneously providing a large number of crystallization environments. Control over changed solution conditions may result from a variety of techniques, including but not limited to metering volumes of crystallizing agent into the chamber by volume exclusion, by entrapment of volumes of crystallizing agent determined by the dimensions of the microfabricated structure, or by cross-channel injection of sample and crystallizing agent into an array of junctions defined by intersecting orthogonal flow channels.
摘要:
The present invention relates to microfluidic devices and methods facilitating the growth and analysis of crystallized materials such as proteins. In accordance with one embodiment, a crystal growth architecture is separated by a permeable membrane from an adjacent well having a much larger volume. The well may be configured to contain a fluid having an identity and concentration similar to the solvent and crystallizing agent employed in crystal growth, with diffusion across the membrane stabilizing that process. Alternatively, the well may be configured to contain a fluid having an identity calculated to affect the crystallization process. In accordance with the still other embodiment, the well may be configured to contain a material such as a cryo-protectant, which is useful in protecting the crystalline material once formed.
摘要:
High throughput screening of crystallization of a target material is accomplished by simultaneously introducing a solution of the target material into a plurality of chambers of a microfabricated fluidic device. The microfabricated fluidic device is then manipulated to vary the solution condition in the chambers, thereby simultaneously providing a large number of crystallization environments. Control over changed solution conditions may result from a variety of techniques, including but not limited to metering volumes of crystallizing agent into the chamber by volume exclusion, by entrapment of volumes of crystallizing agent determined by the dimensions of the microfabricated structure, or by cross-channel injection of sample and crystallizing agent into an array of junctions defined by intersecting orthogonal flow channels.
摘要:
High throughput screening of crystallization of a target material is accomplished by simultaneously introducing a solution of the target material into a plurality of chambers of a microfabricated fluidic device. The microfabricated fluidic device is then manipulated to vary the solution condition in the chambers, thereby simultaneously providing a large number of crystallization environments. Control over changed solution conditions may result from a variety of techniques, including but not limited to metering volumes of crystallizing agent into the chamber by volume exclusion, by entrapment of volumes of crystallizing agent determined by the dimensions of the microfabricated structure, or by cross-channel injection of sample and crystallizing agent into an array of junctions defined by intersecting orthogonal flow channels.
摘要:
High throughput screening of crystallization of a target material is accomplished by simultaneously introducing a solution of the target material into a plurality of chambers of a microfabricated fluidic device. The microfabricated fluidic device is then manipulated to vary the solution condition in the chambers, thereby simultaneously providing a large number of crystallization environments. Control over changed solution conditions may result from a variety of techniques, including but not limited to metering volumes of crystallizing agent into the chamber by volume exclusion, by entrapment of volumes of crystallizing agent determined by the dimensions of the microfabricated structure, or by cross-channel injection of sample and crystallizing agent into an array of junctions defined by intersecting orthogonal flow channels.
摘要:
The present methods are exemplified by a process in which maternal blood containing fetal DNA is diluted to a nominal value of approximately 0.5 genome equivalent of DNA per reaction sample. Digital PCR is then be used to detect aneuploidy, such as the trisomy that causes Down Syndrome. Since aneuploidies do not present a mutational change in sequence, and are merely a change in the number of chromosomes, it has not been possible to detect them in a fetus without resorting to invasive techniques such as amniocentesis or chorionic villi sampling. Digital amplification allows the detection of aneuploidy using massively parallel amplification and detection methods, examining, e.g., 10,000 genome equivalents.