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Table of Contents
1. ISFET Array for Detecting a Single Nucleotide Polymorphism
Priority: Mar 30, 2012
(from WO 2013/144580 A1)
1.1 Technical Field
1.2 Background
• 99.9% of the code is similar among the human population and it is the remaining 0.1% that makes the differences.
• Single Nucleotide Polymorphism is a single mutation of a gene that can be found in more than 1% of a population.
• How to detect if a particular sequence is in the genome? If a primer attaches then it matches and it can be extended. This extension of a primer on the target releases pyrophosphate and H+ ions [3]. Therefore, by detecting whether the acidity in the analyte has increased or not, it is possible to determine whether extension has occurred or not; and consequently whether the designed sequence was matched or not.
• …the presently used platform for ISFETs has made them power and memory intensive.
• The presently used platforms are required to process large amounts of data in order to provide a yes/no answer saying a particular polymorphism is detected in the target DNA or not. These intensive requirements are mainly due to the existence of sources of error that need to be cancelled out in the measurement.
1.3 ISFETs in SNP Detection
• The use of an ISFET in sequencing and SNP detection relies on the fact that hydrogen ions are released as a result of hydrolysis of pyrophosphate, a by-product of the extension of primer/probe strands on target DNA strands when they match
• rather than taking an absolute measurement of the pH, detection of its change is enough to be able to determine whether the designed probe with a known sequence matches with the target. Therefore, having a REFET that is used to detect all non-ideal signals and subsequently cancel them out from a working ISFET in a differential configuration, can provide for a reliable monitoring of the pH change [11, 12]
• Consequently, the REFET is exposed to a mixture of non-extendable (i.e. non-matching) primer with nucleotides, enzymes and target DNA strands to provide the same condition as for the working ISFET except the pH change [13].
• FIG. 6 shows the configuration that is typically used during ISFET-based sequencing for SNP detection.
