Ion semiconductor sequencing

Ion semiconductor sequencing is a method of DNA sequencing based on the detection of hydrogen ions that are released during the polymerization of DNA. This is a method of 'sequencing by synthesis', during which a complementary strand is built based on the sequence of a template strand. Ion semiconductor sequencing is a method of DNA sequencing based on the detection of hydrogen ions that are released during the polymerization of DNA. This is a method of 'sequencing by synthesis', during which a complementary strand is built based on the sequence of a template strand. A microwell containing a template DNA strand to be sequenced is flooded with a single species of deoxyribonucleotide triphosphate (dNTP). If the introduced dNTP is complementary to the leading template nucleotide, it is incorporated into the growing complementary strand. This causes the release of a hydrogen ion that triggers an ISFET ion sensor, which indicates that a reaction has occurred. If homopolymer repeats are present in the template sequence, multiple dNTP molecules will be incorporated in a single cycle. This leads to a corresponding number of released hydrogens and a proportionally higher electronic signal. This technology differs from other sequencing technologies in that no modified nucleotides or optics are used. Ion semiconductor sequencing may also be referred to as Ion Torrent sequencing, pH-mediated sequencing, silicon sequencing, or semiconductor sequencing. The technology was licensed from DNA Electronics Ltd, developed by Ion Torrent Systems Inc. and was released in February 2010. Ion Torrent have marketed their machine as a rapid, compact and economical sequencer that can be utilized in a large number of laboratories as a bench top machine. Roche's 454 Life Sciences is partnering with DNA Electronics on the development of a long-read, high-density semiconductor sequencing platform using this technology. In nature, the incorporation of a deoxyribonucleoside triphosphate (dNTP) into a growing DNA strand involves the formation of a covalent bond and the release of pyrophosphate and a positively charged hydrogen ion. A dNTP will only be incorporated if it is complementary to the leading unpaired template nucleotide. Ion semiconductor sequencing exploits these facts by determining if a hydrogen ion is released upon providing a single species of dNTP to the reaction. Microwells on a semiconductor chip that each contain many copies of one single-stranded template DNA molecule to be sequenced and DNA polymerase are sequentially flooded with unmodified A, C, G or T dNTP. If an introduced dNTP is complementary to the next unpaired nucleotide on the template strand it is incorporated into the growing complementary strand by the DNA polymerase. If the introduced dNTP is not complementary there is no incorporation and no biochemical reaction. The hydrogen ion that is released in the reaction changes the pH of the solution, which is detected by an ISFET. The unattached dNTP molecules are washed out before the next cycle when a different dNTP species is introduced. Beneath the layer of microwells is an ion sensitive layer, below which is an ISFET ion sensor. All layers are contained within a CMOS semiconductor chip, similar to that used in the electronics industry. Each chip contains an array of microwells with corresponding ISFET detectors.Each released hydrogen ion then triggers the ISFET ion sensor. The series of electrical pulses transmitted from the chip to a computer is translated into a DNA sequence, with no intermediate signal conversion required. Because nucleotide incorporation events are measured directly by electronics, the use of labeled nucleotides and optical measurements are avoided. Signal processing and DNA assembly can then be carried out in software. The per base accuracy achieved in house by Ion Torrent on the Ion Torrent Ion semiconductor sequencer as of February 2011 was 99.6% based on 50 base reads, with 100 Mb per run. The read-length as of February 2011 was 100 base pairs. The accuracy for homopolymer repeats of 5 repeats in length was 98%. Later releases show a read length of 400 base pairs These figures have not yet been independently verified outside of the company.