Biochemistry 2280A Lecture Notes - Deoxyribonucleoside, Northern Blot, Base Pair

• School: Western University
• Department: Biochemistry
• Course: Biochemistry 2280A
• Professor: Eric Ball

Document Summary

Procedure relies on the enzymatic extension of a primer using the dna of interest as the complementary template done in the presence of base specific chain terminators. A primer (a short oligonucleotide complimentary to the dna of interest) is annealed to the template dna. The normal substrate for dna synthesis have the oh at the 3 position that is the position that gets extended in the polymerization reaction normal site for the next base to be added 2" deoxynucleotide. The key to sequencing is the 2" 3" dideoxynucleoside triphosphate lacks the. 3" hydroxyl it cannot be extended in a polymerization reaction so they act as chain terminators in a polymerization reaction. The 4 reactions are separated by denaturing polyacylamide gel electrophoresis and examined by autoradiography. A similar strategy is used for automated sequencing with the exception that each of the 4 ddntps is tagged with a different fluorescent label, which allows its unique detection.