This is a very very basic question. I've looked at methods such as chromosome sequencing and shotgun sequencing. Wikipedia says that:

DNA sequencing is the process of determining the precise order of nucleotides within a DNA molecule

For instance, with methods such as shotgun, we already have the original DNA. If we have access to the original DNA itself, don't we ALREADY know the sequence of nucleotides?

Why exactly do we need to 're-build' the DNA with methods such as shotgun and chromosome walking, when we already have the original DNA available? If we have the original DNA, don't we already have the original sequence of nucleotides?

It makes me think that these techniques for 'DNA sequencing' is sort-of pointless. But, obviously I am misunderstanding something.

Thanks in advance, any simple pointers would be great.

  • $\begingroup$ In some sense, this is like saying "what is the point of studying a cell, we already have the cell in our hands." We don't know anything about a cell by having a test tube full of them. This goes for DNA, proteins, RNA, etc. as well. You can purify DNA from an organism without knowing the nucleotides. $\endgroup$ – Danny W. Sep 12 '14 at 21:15

No, you don't. Having the DNA in your hand means that you have access to a macromolecule, not more.

Think of the DNA as a book (or a library). From holding the book in your hands, you don't know what the content is. You have to actively read the book to get this information.

Sequencing a piece of DNA is basically the same. Your readout is the sequence of the bases which is essential to understand (or do research on it) a certain piece of DNA. With this knowledge you can see where genes start and end, define introns and exons (for eukaroytes) and so on.

  • $\begingroup$ From a bioinformatics point of view: We are given a string of DNA characters (e.g. CTCGAT), then use shotgun to re-create the exact same string? I don't quite understand the point of that when we have the original string available. $\endgroup$ – lostguy Sep 12 '14 at 23:03
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    $\begingroup$ @lostguy If you're given a DNA sequence, you obviously don't need to sequence it. But what if you're given a DNA molecule where the sequence is unknown? That is where sequencing is useful. $\endgroup$ – canadianer Sep 12 '14 at 23:05
  • $\begingroup$ Oh okay, thanks a lot. that makes more sense. I am doing bioinformatics homework and this type of problem came up. I kept asking myself: "What's the point of this"? But now I see. Out of curiosity, how are methods like shotgun used practically (if we don't have the original DNA string)? Do we physically randomly cut-up DNA? (Hmm, that doesn't seem realistic) $\endgroup$ – lostguy Sep 12 '14 at 23:10
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    $\begingroup$ @lostguy Yes, that's exactly what occurs. DNA is physically sheared into random sized fragments, typically through sonication. Appropriately sized fragments are selected by gel electrophoresis and cloned into plasmids for amplification/sequencing. That's a very general overview, actual experimental protocols will differ and modern (next-gen) sequencing uses different methods. $\endgroup$ – canadianer Sep 12 '14 at 23:19

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