DNA sequencing Flashcards
What is DNA sequencing?
Finding the order of DNA nucleotides or bases in a piece of DNA
2 procedures of DNA sequencing
Selective termination of DNA synthesis at specific nucleotides
High resolution electrophoresis
What is high resolution electrophoresis?
Motion of particles relative to fluid under an electric field
Role of high resolution electrophoresis
Separate fragments of DNA
What is methyl interference assay?
Method to identify nucleotides important for DNA binding
What is produced when polymerase I is cut?
Large protein fragments called Klenow fragments
What cuts DNA polymerase I?
Protease subtilisin enzyme
What is the technique used to separate DNA or protein molecules?
Polyacrylamide gel electrophoresis (PAGE)
What is autoradiography?
X-ray produced by the pattern of decay emissions of a radioactive substance
What is genome sequencing?
Determination of the order of bases in DNA of a genome
What is a genome map?
Set of landmarks in a genome
2 methods of DNA sequencing
Maxam-Gilbert method and Sanger dideoxy method
What is the Maxam-Gilbert method?
Chemical modification of DNA and cleavage at specific bases
Negatives of Maxam-Gilbert method
Complex, uses hazardous chemicals and difficult to scale up
What chemical destroys base G?
Dimethyl sulphate
What chemical destroys base A?
Formic acid
What chemical destroys base T?
Hydrazine
What chemical destroys base C?
Sodium chloride
1st step of Sanger dideoxy method
DNA strands are separated by electrophoresis
2nd step of Sanger dideoxy method
Polyacrylamide gel separates strands
3rd step of Sanger dideoxy method
Autoradiography used to detect radioactive bands
4th step of Sanger dideoxy
Sequence is determined by band on autoradiography
What is high-throughput sequencing?
Lowering cost of DNA sequencing
How do you sequence a genome?
Break genome, sequence the pieces, reassemble in proper order
