4. You perform a PCR reaction using the following template DNA and primers. What

Question

4. You perform a PCR reaction using the following template DNA and primers. What size(s) of products do you expect? Explain vour answer. Primer: 5'-ctctgggctc agagcagccc-3 Primer: 5'-catgcagccg tcctgatctc-3 Template: 1 attatatttt atattatata ctctgggctc agagcagccc 40 41 atattatata tatatatttt aaaatattat aaatttattt 80 81 catgcagccg tcctgatctc attatatttt ataatttttt 120 121 ttttattttt attatatttt ttttttattt ttattttttt 160 161 aaaatattat tatattatta taattaattt attataaaat 200 201 tatatatttt attatatttt tatatatttt attaaatttt 240 241 attatatttt tttttaattt ttattattat tttttatata 280 281 tatatatttt tatatatttt attatatttt attatatttt 320 321 tttatataaa attatatttt tatttttttt ttttttataa 360 361 gggctgctct gagcccagag attatatttt attataaaat 400 401 attatatttt attataaaat aaaatataat aaaatattat 440 441 aaaatattat aaaaaaaaat gagatcagga cggctgcatg 480 481 atattatata atattatata attatatttt atattatata 500

Explanation / Answer

Polymerase chain reaction (PCR) used to amplify the DNA in a cell-free system and it is alternate for gene cloning. It is first discovered by the Kary Mullis in 1983 and he awarded the noble prize in 1993. The requirements for PCR are genomic DNA, a forward primer and reverse primer, PCR buffer, dNTPs, Taq polymerase and deionized water.

Template Strand

ATTATATTTTATATTATATACTCTGGGCTCAGAGCAGCCCATATTATATATATATATTTTAAAATATTATAAATTTATTTCATGCAGCCGTCCTGATCTCATTATATTTTATAATTTTTTTTTTATTTTTATTATATTTTTTTTTTATTTTTATTTTTTTAAAATATTATTATATTATTATAATTAATTTATTATAAAATTATATATTTTATTATATTTTTATATATTTTATTAAATTTTATTATATTTTTTTTTAATTTTTATTATTATTTTTTATATATATATATTTTTATATATTTTATTATATTTTATTATATTTTTTTATATAAAATTATATTTTTATTTTTTTTTTTTTTATAAGGGCTGCTCTGAGCCCAGAGATTATATTTTATTATAAAATATTATATTTTATTATAAAATAAAATATAATAAAATATTATAAAATATTATAAAAAAAAATGAGATCAGGACGGCTGCATGATATTATATAATATTATATAATTATATTTTATATTATATA                       

Forward primer        5’-CTCTGGGCTCAGAGCAGCCC-3’

Reverse primer          5’-CATGCAGCCGTCCTGATCTC-3’

After the binding of primers (the primer binding sites on template strand are represented by the underline & italic portions) the Taq polymerase extends the 3’ hydroxyl groups of primes. The product produced after PCR amplification is 460 base pairs (middle portion between the primer binding sites on template).

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