Question

Based on the following picture:

1- Write the DNA sequence of the recombinant pET/GFP plasmid beginning at the ATG translational start codon and continuing through the first 4 codons (triplets) of the GFP coding region. Put a box around the NheI site.1-

2- Also, based on the DNA sequence you have written out above, write the amino acid sequence of the protein that is encoded by this DNA. Box the His-tag. Underline the region that corresponds to the GFP protein sequence.

Indicate the start site of transcription from the region of the pET28 vector drawn above, indicate the position of the translational start codon, and indicate the position where did cut this vector to insert DNA that contains the coding region of GFP?

Tarometer T7 promoter Lac operator la creator o Xbal r as | History RBS His-tag Nhel Model EcoRI

Answer 1

T7 PROMOTER LAC OPERATOR XBA1 RBS

TTAATACGACTCACTATAGGGGAATTGTGAGCGGATAACAATTCCCCTCTAGAAATAATTTTGTTTAACTTTAAGAA

HISTAG NHE1

GGAGA TATACCATGGGCAGCAGCCATCATCATCATCATCAGCTAGCATGGTGAGCAAGGGCGAGGAGCTGTTCACCGGGGTGGTGCCCATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGCGAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTGCCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGCTACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTCCAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAGTTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGACGGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATGGCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGACGGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTGCTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAGAAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATGGACGAGCTGTACAAGTAAGAATTC

ECOR1

HISTAG GFP

2.MGSSHHHHHASMVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYKQF

3.T7 PROMOTER is the transcription start site ...and MGSS is the translation start site .To clone the dna in PET2