How does 454 pyrosequencing work

The method used in this sequencing process is pyrosequencing. Put a smaller bead into the nanopore, and start the sequencing reaction. Finally the results are processed by computer software. Because each kind of dNTP produces unique fluorescence color in the reaction, DNA sequence can be measured according to the fluorescence colors.

After the reaction, ATP are degraded by diphosphatase, leading to fluorescence quenching, so that sequencing reaction goes into the next cycle. For sequencing technology, each reaction is carried out in independent nanopore of PTP board, thus it greatly reduces mutual interference and sequencing bias.

The biggest advantage of technology is to get longer sequencing read length. Currently, average read length of technology is up to bp. The disadvantage of is that it is unable to accurately measure the homopolymer length. For this unavoidable reason, technology will introduce insertion and deletion sequencing errors to the results. Types of genome sequencing. DNA Sequencing. How helpful was this page?

What's the main reason for your rating? Which of these best describes your occupation? What is the first part of your school's postcode? How has the site influenced you or others? Thankyou, we value your feedback! We use cookies to improve this site. Overview [ ] pyrosequencing is a method of high throughput DNA sequencing that utilizes a single strand of DNA with a length of bp. Luciferase: Uses ATP to convert luciferin to oxyluciferin, which is a molecule that emits visible light.

Apyrase: Degrades unincorporated nucleotides. The oxyluciferin emits light that can be captured with a very sensitive c Pyrosequencing sequence output chart. QIAGEN amera and analyzed in a program to produce a graph showing when light was captured and how much light was captured in relation to which dNTP was added to the incubation.

If a dNTP does not match, no light is emitted. References: Pyrosequencing animation. Technology page.