Practice problem I have performed PCR using primers for the Cox-1 gene in Tardig

Question

Practice problem I have performed PCR using primers for the Cox-1 gene in Tardigrades, I cloned my PCR product into the TOPO cloning vector, and now I have to sequence it to determine the species of Tardigrade. I have given you the sequence of the TOPO cloning vector and the primer used to start the sequencing reaction. Show me what you would find in the ddNTP tubes and draw out the sequencing gel. 5' ccgggttattcccggtta 3' The primer used for the sequencing reaction is 5' taacc 3' 1. E (2 pts): draw gel A (2 pts): ddGTP tube: B (2 pts): ddATP tube: C (2 pts): ddTTP tube: D (2 pts): ddCTP tube:

Explanation / Answer

Different fragments of DNA molecules that are obtained after sequencing the fragments with Sanger dideoxymethod is as follows-

a) ddGTP: 5'-TAACCG-3' , 5'-TAACCGG -3' , 5'-TAACCGGG -3' , 5'-TAACCGGGAATAACCCG-3' ,    5'-TAACCGGGAATAACCCGG-3'

b) ddATP: 5'-TA-3' , 5'-TAA-3' , 5'-TAACCGGGA-3' , 5'-TAACCGGGAA-3' , 5'-TAACCGGGAATA-3' ,    5'-TAACCGGGAATAA-3'

c) ddTTP: 5'-T-3' , 5'-TAACCGGGAAT-3'.

d) ddCTP: 5'-TAAC-3' , 5'-TAACC-3' , 5'-TAACCGGGAATAAC-3' , 5'-TAACCGGGAATAACC-3' , 5'-TAACCGGGAATAACCC-3'   

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