Biochemistry 2280A Lecture Notes - 2Degrees, Cell Nucleus, Formamide
20 Nov 2012
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Case study: you have identified a human protein that will be of tremendous value in treating disease. Your goal is to mass produce this protein (yfp) Obtaining yfp will require: cloning your favorite gene, introducing yfg into e. coli, purifying the protein from e. coli. How do we get enough of yfg to be able to clone it: synthetic oligonucleotides, hybridization, pcr. Synthetic oligonucleotides: short fragments (20-100 bases) of single stranded dna will defined sequence can be made synthetically and inexpensively, often referred to as primers or oligos. Nucleic acid hybridization: denaturation/melting, renaturation/annealing/hybridization, dna denaturation: if you heat a double strand dna or put it into a high ph, the strands will separate into single stands. Melting temperature: given a specific dna at a concentration of 1 m, the temperature at which 50% of the molecules are single stranded (referred to as the tm , every different dna molecule has a different tm.
