BIOL10003 Lecture Notes - Lecture 33: Gel Electrophoresis, Polymerase Chain Reaction, Dna Ligase

• School: University of Melbourne
• Department: Biology
• Course: BIOL10003
• Professor: All

Document Summary

Denaturing/melting melting h+ bonds, breaks hydrophobic nucleobases, temperature (80 +), chemical (alkali) Renaturing (dna-dna)/hybridisation (dna-rna) low temperature (<80 c), neutral ph. Oligonucleotide primers - start and stop of the region (in excess) Deoxyribonucleotide triphosphates (dntp) - building blocks (in excess) Steps (cycled and repeated by changing temperature): denaturation (dsdna) 95 c, annealing/hybridisation (primers) 45-60 c, extension (dna polymerase) 70 c. Bacteria dna is protected; methyl groups at recognition sites. Different types (i-v): type ii is important for this course. Type ii enzymes cut within or close by this sequence; (cid:271)lunt, sti(cid:272)ky (cid:894)3" or 5" overhang(cid:895) Isoschizomers (can be joined) are pairs of restriction enzymes specific to the same recognition sequence. Neoschizomers recognize the same nucleotide sequence as their prototype but cleave at a different site. Dna is negatively charged due to negative phosphate group and will migrate from -ve towards. Thickness of gel - thicker = slower.