You want to express a rare human protein in bacteria so that you can make large

Question

You want to express a rare human protein in bacteria so that you can make large quantities of it. To aid in its purification, you decide to add a stretch of six histidines to the N-terminus or the C-terminus of the protein. Such histidine-tagged proteins bind tightly to Ni2+ columns but can be readily eluted with a solution of EDTA or imidazole. This procedure allows an enormous purification in one step. The nucleotide sequence that encodes your protein is shown in the figure below. Design a pair of PCR primers, each with 18 nucleotides of homology to the gene, that will amplify the coding sequence and add an initiation codon followed by six histidine codons to the N-terminus. Design a pair of primers that will add six histidine codons followed by a stop codon to the C-terminus.

Explanation / Answer

PCR primers each with 18 nucleotides

Forward primer = 5' GAA CGA CGT TCA GAG AGA 3'

Reverse primer = 3' ACG AGT AGC CAT ACG ACC 5'

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