BCMB20005 Lecture Notes - Lecture 4: Taq Polymerase, Molecular Cloning, Complementary Dna

• School: University of Melbourne
• Department: Biochemistry and Molecular Biology
• Course: BCMB20005
• Professor: Amber Willems

Document Summary

All figures from lecture 4 slides, a willems-jones, march 2017, melbourne university (unless stated) Pcr: principle, mechanism and applications, modern cloning using pcr compared to traditional cloning strategy. Rt-pcr: cdna: what is it and how is it generated, process of rt-pcr. Quantitative/real time pcr: two types, principle. Developed in 1983 revolutionised molecular biology and cloning. Pcr: in vitro amplification of a dna sequence: exponential copying of few dna molecules into thousands-millions of copies. Utilizi(cid:374)g (cid:862)(cid:373)elti(cid:374)g(cid:863) a(cid:374)d (cid:862)a(cid:374)(cid:374)eali(cid:374)g(cid:863) p(cid:396)ope(cid:396)ties of dna. Requires some knowledge of the sequence of the gene of interest for primers: sequence specific primers hybridize complementary dna. Paternity testing: genetic markers, looking at variation between people. Dna pol(cid:455)(cid:373)e(cid:396)ase, e. g. ta(cid:395) pol(cid:455)(cid:373)e(cid:396)ase: (cid:272)atal(cid:455)se step(cid:449)ise additio(cid:374) of (cid:374)u(cid:272)leotides at (cid:1007)" e(cid:374)d: requires a pre-existing strand to add these nucleotides, dna is e(cid:454)te(cid:374)ded i(cid:374) a (cid:1009)" to (cid:1007)" di(cid:396)e(cid:272)tio(cid:374) Sequence-specific primers (oligonucleotides) must flank dna sequence needed for : forward and reverse primer. Nucleotide bases (dntps): datp, dctp, dttp, dgtp.