Genomics Wonwoo Jeong
1. Genomics
1) Definition
Analyzing function and structure of genomes discipline. Using DNA sequencing. Gilbert and Sanger method is used to verify DNA sequence that cleavage DNA strand using ddNTPs and dNTPs.<o:p></o:p>
2) Application
Human Genome Project is the most famous application of genomics. The project is clarifying sequence of chemical base pairs. Not only human but also any other animals can be subjected to this project. By mapping organism’s genome in a row, it is possible to figure out taxonomical relationship as well. Relationship between primates and humankind, between Siberia tiger and Bengal tiger and between any other co-existent codes.<o:p></o:p>
3) Sanger method
(http://en.wikipedia.org/wiki/Sanger_sequencing)<o:p></o:p>
The classical chain-termination method requires a single-stranded DNA template, a DNA primer, a DNA polymerase, normal deoxynucleotidetriphosphates (dNTPs), and modified di-deoxynucleotidetriphosphates (ddNTPs), the latter of which terminate DNA strand elongation. These chain-terminating nucleotides lack a 3'-OH group required for the formation of a phosphodiester bond between two nucleotides, causing DNA polymerase to cease extension of DNA when a modified ddNTP is incorporated. The ddNTPs may be radioactively or fluorescently labeled for detection in automated sequencing machines.<o:p></o:p>
The DNA sample is divided into four separate sequencing reactions, containing all four of the standard deoxynucleotides (dATP, dGTP, dCTP and dTTP) and the DNA polymerase. To each reaction is added only one of the four dideoxynucleotides (ddATP, ddGTP, ddCTP, or ddTTP), while the four other nucleotides are ordinary ones. The dideoxynucleotide is added in approximately 100-fold excess of the corresponding deoxynucleotide(e.g. 0.5mM ddATP : 0.005mM dATP) allowing for enough fragments to be produced while still transcribing the complete sequence.[2] Putting it in a more sensible order, four separate reactions are needed in this process to test all four ddNTPs. Following rounds of template DNA extension from the bound primer, the resulting DNA fragments are heat denatured and separated by size using gel electrophoresis. In the original publication of 1977,[2] the formation of base-paired loops of ssDNA was a cause of serious difficulty in resolving bands at some locations. This is frequently performed using a denaturing polyacrylamide-urea gel with each of the four reactions run in one of four individual lanes (lanes A, T, G, C). The DNA bands may then be visualized by autoradiography or UV light and the DNA sequence can be directly read off the X-ray film or gel image. (http://en.wikipedia.org/wiki/Sanger_sequencing)<o:p></o:p>
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