1. Given the sequence for the cDNA of human Beta globin, make PCR primers that w

Question

1. Given the sequence for the cDNA of human Beta globin, make PCR primers that would allow you to clone it into the GFP-C1 vector using Kpnl (5' end) and BamHI (3' end). Put restriction enzyme sequence in the BOLD blanks and fill the remaining 10 blanks with sequence complimentary to beta globin. (8 pts) Beta globin coding sequence 5' ATGGTGCACCTGACTCCTGAGGAGAAGTCTGCCGTTACTGCCCTGTGGGGCAAGGTGAACGTGGATGAAGTT GGTGGTGAGGCCC TGGGCAGGCTGC TGGT GGTCTACCC TTGGACCCAGAGGTTCT TTGAGTCC TT TGGGGATCT GTCCACCCCTGATGCTGTTATGGGCAACCCTAAGGTGAAGGCTCATGGCAAGAAAGTGCTCGGTGCCTTTAGTG ATGGCCTGGCTCACCTGGACAACCTCAAGGGCACCTTTGCCACACTGAGTGAGCTGCACTGTGACAAGCTGCAC GTGGATCCTGAGAACTTCAGGCTCCTGGGCAACGTGCTGGTCTGTGTGCTGGACCATCACTTTGGCAAAGAATT CACCCCACCAGTGCAGGCTGCCTATCAGAAAGTGGTGGCTGGTGTGGCTAATGCCCTGGCCCACAAGTATCACT AC 3 Beta globin protein sequence (not needed for this lab-just for reference) MVHLTPEEKSAVTALWGKVNVDEVGGEALGRLLVVYPWTOREFESFGDLSTPDAVMGNPKVKAHGKKVLGAFSD GLAHLDNLKGTFATLSELHCDKLHVDPENFRLLGNVLVCVLDHHFGKEFTPPVQAAYQKVVAGVANALAHKYH

Explanation / Answer

Restriction site of Kpn1 = GGTACC

Restriction site of Bam H1 = GGATCC

Forward Primer

5'-GGTACCATGGTGCACC-3'

Reverse Primer

Reverse and complement of 5'-GTATCACTAC-3' is

5'-GTAGTGATAC-3'

Therefore, the reverse primer is:

5'-GGATCCGTAGTGATAC-3'

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