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Introduction to Oxford Nanopore TechnologiesMinIon

Oxford Nanopore Technologies developed a new generation of nanopore-based electronic systems for analysis of single molecules, including DNA, RNA and proteins.
The characteristics of the technology is low costs, the technology is portable, the data can be generated in real time and no amplification step of the target DNA is needed prior sequencing. Currently, the technology is still in an extensive testing phase, thus limited published data are available. Compared to other NGS technologies the Oxford Nanopore Technologies system still displays a higher sequencing error rate, the base calling accuracy is indicated to be up to 96%.

How does Oxford Nanopore Technologies sequencing works? NanoporeSeqTechnology

The Oxford Nanopore Technologies is based on protein nanopores that are inserted in membranes created by synthetic polymers. Nanopores are nano-scale holes formed by proteins or synthetic materials. Each membrane contains thousands of such nanopores. An electronic potential is applied across the membrane, if molecules or other analytes flow through the nanopore a characteristic disruption in the current of the nanopore can be measured. This characteristic current changes allows to distinguish between the four standard DNA bases (G, A, T, and C) and enables sequencing of DNA strands if they are passing the nanopore. During the sequencing process an enzyme/ DNA template complex is approaching the nanopore. The enzyme unzips the double stranded DNA in single strands which it guides through the nanopore one base at a time. During this process the characteristic disruptions in current will be measured for each individual nucleotide passing the nanopore. The specific sequence of measured current change signals will be translated into the DNA sequence information. In order to improve the accuracy the DNA template bound to the enzyme will contain a hairpin structure at one end. This hairpin structure prevents the complete cleaving of the two DNA single strands, and the second DNA strand will also pass through the nanopore. Thus the system can sequence both strands the sense and the antisense DNA strand and combine these information, this improves data analysis and base calling accuracy.
The following video shows in more detail how nanopore technology works.


Oxford Nanpore Technologies sequencing time, read length and throughput

The sequencing speed is 280-500 base pairs per second. As the technology enables real time sequencing first data will be available after two minutes. The maximal (current default) sequencing run time is 48 hours.
The read length correspond to the DNA fragment length. The longest reported read length is between 230 and 300 kilo base pairs. The average read length depends on used fragmentation methods.
The estimated number of reads (10 kilo base length) is between 2.5 and 4.4 million for the MinION™, 14.5-26 million for the PromethION™, and 700 to 1250 million for the GridION™ system.


Oxford Nanpore Technologies sequencing devices and its applicationsGridION

Oxford Nanpore Technologies developed different sequencing devices, the handheld MinION™ and the high-throughput/high sample number PromethION™ and GridION™ systems.
The MinION is commercially available. The PromethION is currently tested in an Early Access programme (PEAP).
As this technology enables sequencing of RNA no transcription into DNA is required. This can be beneficial for sequencing of certain RNA molecules such as viral RNA genomes. PromethIONFurthermore, the technology can be applied if long reads are especially beneficial such as for sequencing complex regions with structural variants including insertions, deletions, duplications, recombination, inversions or repetitive elements
This sequencing technology may be used in a number of applications in scientific research, personalised medicine, crop science, security and defence and environmental studies.
For more information about sequencing systems and its application visit the Oxford Nanopore Technologies website.