Press the start button then select the most appropriate word to fill each gap from the drop-down list. Each word in the list is used only once. Hint: read ahead and think about grammar to narrow down the choice of possible answers. When you think that you have it right, press the check sequence button. There is a 5 second penalty if you are wrong!
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DNA sequencing - the ability to determine the exact order of in a molecule of DNA - has triggered a revolution in our understanding of biology. The technology was developed in the 1970s, building on earlier methods developed to sequence and short sequences of RNA. The original method relied on three key developments. The ability to tag one end of a DNA molecule; chemical reactions that DNA after specific bases; and high resolution that was capable of separating DNA fragments that differed in size by only one base.
This approach, coupled with the ability to clone genes of interest in bacterial through the use of enzymes and , enabled scientists to sequence a large number of genes. It was overtaken, however, by another approach that avoided chemical cleavage that was developed by Fred Sanger in Cambridge.
Instead of fragmentation of intact DNA, sequencing relies on to generate a DNA copy of the region of interest. The method uses a reaction mixture with low concentrations of in addition to the normal . Dideoxynucleotides can be incorporated into the growing DNA chain as normal, but the lack of a 3' group on the nucleotide sugar means that the chain cannot be extended, and a chain is released. The development of dideoxy terminators that carried specific dye molecules meant that it became possible to generate a set of labelled DNA fragments from the target sequence in a single tube. A complete set of fragments differing in length by one base is generated, each labelled with a dye that defines the identity of the base at that position.
This improved approach led to the development of sequencers, capable of sequencing tens of thousand of bases in a single run. In combination with the ability to large regions of DNA in specialised DNA such as BACs (bacterial artificial chromosomes) and new strategies such as sequencing (sequencing random fragments of DNA and relying on computers to assemble the overall sequence), it opened the way to sequencing entire genomes, leading to the first draft of the 3 billion bp human genome sequence in 2000.
Automated dideoxy sequencing is still in widespread use, but for sequence analysis it has been superseded by so-called NGS ( sequencing approaches). The most widely used at the moment is based on imaging millions of DNA chains as they grow on a support, using fluorescent dyes and computer imaging to collect the data. These sequencers collect millions of short DNA sequences in 24 hr, and use fast to stitch these together using sequence overlaps. The first human genome sequence was a major international effort that took more than three years and cost several hundred million dollars. Next generation sequencing has reduced the time needed to sequence a human genome to under a week, and reduced the cost to about £1000. This means that routine genome sequencing is a practical possibility, though the challenge is to work out the best way to use the data to improve care.