Biology 1002B Lecture Notes - Lecture 20: Sanger Sequencing, Dideoxynucleotide, Pyrosequencing
30 Mar 2016
School
Department
Course
Professor
Document Summary
Dna sequences methods typically have these basic steps. Dna replication is not done in one continuous strand. Similarly, we do not have the technology to sequence large genome is one continuous strand (can"t sequence big genome all in once) Must be broken up into pieces/fragments and sequence each fragment. Must put together the fragments into one strand. Get sequence as you add on bases of the complementary strand. Knows the sequence has an a because dna polymerase wanted to add a t. 4 separate tubes each containing a mixture of regular deoxyribonucleotide (99%) and one type of the dideoxynucleotide (1%) Dideoxynucleotide bases lack a 3" -oh end. Elongation stops when dideoxynucleotide is added (terminate synthesis) Depends when the dideoxynucleotide was incorporated by chance. Most of the time, a normal base will be added but about 1% of the time, a dideoxynucleotide is added. The fragment are sorted by size by electrophoresis.
