Biology > Lab Report > Hunter College, CUNYBIO 203Biology 203 LAb2 (All)

Hunter College, CUNYBIO 203Biology 203 LAb2

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Lab Report II (Biol 203, Spring 2019) NAME (First, LAST): Kabongo, Charlene Sessions 9,10 PCR Cloning and Expression of a GFP fusion protein in E. coli PART I (No lab data required)- 60 points 1. ... Below is a map for a plasmid called ‘pGFP’: The sequence (5’ to 3’) of pGFP containing the promoter region, the 5’ multiple cloning site (with a BamHI site, underlined), and the 5’ end of the GFP coding sequence is as follows: 1 AGCGCCCAAT ACGCAAACCG CCTCTCCCCG CGCGTTGGCC GATTCATTAA TGCAGCTGGC 61 ACGACAGGTT TCCCGACTGG AAAGCGGGCA GTGAGCGCAA CGCAATTAAT GTGAGTTAGC 121 TCACTCATTA GGCACCCCAG GCTTTACACT TTATGCTTCC GGCTCGTATG TTGTGTGGAA 181 TTGTGAGCGG ATAACAATTT CACACAGGAA ACAGCTATGA CCATGATTAC GCCAAGCTTG 241 CATGCCTGCA GGTCGACTCT AGAGGATCCC CGGGTACCGG TAGAAAAAAT GAGTAAAGGA 301 GAAGAACTTT TCACTGGAGT TGTCCCAATT CTTGTTGAAT TAGATGGTGA TGTTAATGGG 361 CACAAATTTT CTGTCAGTGG AGAGGGTGAA GGTGATGCAA CATACGGAAA ACTTACCCTT 1The GFP open reading frame was cloned as a BamHI (5’ MCS)/ XhoI (3’MCS) fragment. In this plasmid, GFP is expressed as a fusion protein with LacZ. Clearly indicate on the above sequence the following elements: lac Promoter : lacP The start codon for LacZ: LacZ#1 The first codon of the GFP sequence : GFP#1 (here are the first ten amino acids for GFP: MSKGEELFTG) Lac Operator : lacO The transcription start (+1) of the mRNA : circle the nucleotide and indicate +1 The Shine Dalgarno sequence : SD Is the plasmid-encoded fusion protein subject to regulation by cAMP (catabolite repression)? Explain your answer. HINT: To do this, you will need to consult figures 12. [Show More]

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