Discussion of all aspects of biological molecules, biochemical processes and laboratory procedures in the field.
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So I understand that fluorescent markers are attached to each of the dideoxy nucleotides and then the DNA fragments are aligned and read to determine their sequence.. why not attach fluorescent markers to the regular nucleotides and then be able to read the sequence straight from there?
How will you separate out the color signals if they are all mixed together? Even if you could sort them spectrally and learn the relative amount of each color fluor present, how would you know where in the sequence they are in along the DNA molecule? Terminating a growing chain with a single dideoxy allows you to run the mixture of different chain lengths out on a gel, sorting by size, with a different pure color at the end of each sized molecule. That way you simplify the signal (single color) and sort the molecules by size so you can read off the sequence as colors arranged along the gel lane.
Last edited by jonmoulton on Wed Mar 03, 2010 4:01 pm, edited 1 time in total.
Okay there's something I'm not understanding here and I'll probably feel really stupid in a minute. I thought the dideoxy nucleotides were tagged with different colours you could see under a microscope- so then I thought, why not tag the regular nucleotides and look at the colour sequence from there?
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