Applied Biosystems

About: Applied Biosystems is a based out in . It is known for research contribution in the topics: Mass spectrometry & Nucleic acid. The organization has 1521 authors who have published 1579 publications receiving 285423 citations.

System and method for preparing and using bulk emulsion

Abstract: An emulsion generation apparatus and method for forming an emulsion are provided wherein a customized impeller design is adapted to form an emulsion with a desired droplet size that defines a desired volume. The emulsion generation apparatus provides improved uniformity in emulsion preparation and may be used to create large or small volume emulsions rapidly and reproducibly. A system and method are also provided for large volume sample amplification adaptable for use with conventional PCR-based reactions as well as emulsion-based PCR reactions and other reactions. For applications involving emulsion- based PCR amplification, the system and method provide improved uniformity in emulsion amplification and can be used to amplify large or small volume emulsions rapidly and reproducibly.

Asynchronous primed pcr

Abstract: An asynchronous thermal cycling protocol for nucleic acid amplification uses two primers with thermal melting temperatures different by about 10 to 30 °C. After the higher melting primer has annealed and polymerase mediated extension, the uncopied, single-stranded target sequence may be hybridized and detected by a probe. DNA probes may be cleaved by the exonuclease activity of a polymerase. The probe may be a non-cleaving analog such as PNA. When a probe is labelled with a reporter dye and a quencher selected to undergo energy transfer, e.g. FRET, fluorescence from the reporter dye may be effectively quenched when the probe is unbound. Upon hybridization of the probe to complementary target or upon cleavage while bound to target, the reporter dye is no longer quenched, resulting in a detectable amount of fluorescence. The second, lower-melting primer may be annealed and extended to generate a double-stranded nucleic acid. Amplification may be monitored in real time, including each cycle, or at the end point. The asynchronous PCR thermal cycling protocol can generate a preponderance of the PCR amplicon in single-stranded form by repetition at the end of the protocol of annealing and extension of the higher melting primer.

A one-step coupled amplification and oligonucleotide ligation procedure for multiplex genetic typing

Abstract: A new technique, coupled amplification and oligonucleotide ligation (CAL), has been developed that allows for simultaneous multiplex amplification and genotyping of DNA. CAL is a biphasic method that combines in one assay DNA amplification by PCR with DNA genotyping by the oligonucleotide ligation assay (OLA). By virtue of a difference in the melting temperatures of PCR primer-target DNA and OLA probe-target DNA hybrids, the method allows preferential amplification of DNA during stage I and oligonucleotide ligation during stage II of the reaction. In stage I, target DNA is amplified using high-melting primers (Tm values between 68 degrees C and 89 degrees C) in a two-step PCR cycle that employs a 94 degrees C denaturation step and a 72 degrees C anneal-elongation step. In stage II, genotyping of PCR products by competitive oligonucleotide ligation with oligonucleotide probes (Tm values between 51 degrees C and 67 degrees C) located between the PCR primers is accomplished by several cycles of denaturation at 94 degrees C followed by anneal-ligation at 55 degrees C. Ligation products are fluorochrome-labeled at their 3' ends and analyzed electrophoretically on a fluorescent DNA sequencer. The CAL procedure has been used successfully to analyze human genomic DNA for cystic fibrosis (CF) alleles. Because product detection occurs concurrently with target amplification, the technique is rapid, highly sensitive, and specific and requires minimal sample processing.

Whole genome expression analysis system

Abstract: A method for simultaneously determining a genetic expression profile for an individual member of a species relative to an entire standard genome for the species. The method can comprise distributing a liquid sample into an array of reaction chambers of a substrate. The array can comprise a primer set and a probe for each polynucleotide target along the entire standard genome. The liquid sample can comprise substantially all genetic material of the member. Each of the reaction chambers can comprise the primer set and the probe for at least one of the polynucleotide targets and a polymerase. The method can further comprise amplifying the liquid sample in the array, detecting a signal emitted by at least one of the probes, and identifying the genetic expression profile in response to the signal.