Biotech Techniques

Vector Cloning:
- natural DNA replication
- restriction sites
- incomplete digestion: not all sites are cut (no money no time)
- complete digestion: all sites are cut (have money have time)
- restriction enzymes cut covalent phosphodiester bonds of both strand
- DNA fusions can be name permanent by DNA ligase (seals strand by catalyzing formation of phosphodiester bonds)
- restriction enzymes need sticky ends
- fragment placed into plasmid of bacterial cell (transformation)
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PCR (Polymerase Chain Reaction):
- need DNA template, dNTP (dATP, dCTP, dGTP, dTTP), primers (need TWO known sequence), Taq polymerase (heat resistant)
- don't need primase, helicase, gyrase, ligase, and ssbps because it uses heat instead

Sanger's Method of Sequencing:
- need: DNA template, dNTP (dATP, dCTP, dGTP, dTTP), primers (need ONE known sequence), polymerase
- don't need primase, helicase, gyrase, ligase, and ssbps because it doesn't use heat
- need ddNTP (dideoxy NTP, two no oxygens), signals the stop of elongation since there is no oxygen for anything to attach on
Sequencing:
- the whole strand is a possibility
- strand can we cut on any letter depending on the ddNTP

Vector Cloning and PCR:
- used to replicate specific gene of interest

PCR and Sanger's Sequencing:
- need parental/ template strand
- artificial DNA replication

Vector Cloning, PCR, Sanger's Sequencing:
- DNA replicated and made in mass production