Answer Happy

In lab, you amplified the GFP coding sequence by PCR as an Xhoſ fragment, in order to clone it into the Xhol site of the vector PSK. The GFP insert was amplified in such a way that, after ligation, the GFP coding sequence was in frame with LacZá present on the plasmid, thereby producing a construct expressing a Lacza-GFP fusion protein in E. coli. Your task is to design the 5' oligonucleotide used to amplify GFP in order to obtain the correct PCR product to clone into pSK (assume you have the 3' oligonucleotide available). Your 5' oligonucleotide needs an Xhol site, which, after restriction, will produce an overhang that can be ligated in frame into the pSK vector. You will need to write down the sequence of your 5' oligonucleotide as follows: 1) the restriction site (6 nucleotides) Ans: 2) Any nucleotide you may find necessary to include to preserve the frame (0,1 or 2 nucleotides) Ans: 3) 5 codons (15 nucleotides) corresponding to the relevant GFP coding sequence, starting from the second codon of GFP. Ans: