Automated DNA Sequencing Flashcards Preview

OCR Biology F215 > Automated DNA Sequencing > Flashcards

Flashcards in Automated DNA Sequencing Deck (2)
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1
Q

What are the steps involved in automated DNA sequencing?

A
  • the primer anneals to the 3’ end of the template strand, allowing DNA polymerase to attach
  • DNA polymerase adds free nucleotides according to base pairing rules so the strand grows
  • if a modified nucleotide (terminator base) is added the polymerase enzyme is thrown off and the reaction stops on that template strand
  • as the reaction proceeds many molecules of DNA are made and the fragments vary in size, the final added nucleotide of each strand is tagged with a specific colour
  • the fragments are then run by electrophoresis to separate the fragments by size
  • the strands are then run through a machine and a laser reads the sequence in order of size from the bottom to the top
  • there is a sequence of colours displayed so the DNA sequence can be built up one base at a time
2
Q

What is included in the reaction mixture?

A
  • DNA polymerase
  • many copies of the single stranded DNA fragment
  • free nucleotides
  • primers
  • fluorescently labelled modified nucleotide - if they are added to the growing chain the DNA polymerase is ‘thrown off’ and the strand cannot have any further nucleotides added to it, each different nucleotide has a different coloured fluorescent marker attached to it