Molecular Biology, PCR homework, need help. For all primers, use 15 bp of comple

Question

Molecular Biology, PCR homework, need help.

For all primers, use 15 bp of complementarity to the target DNA. 1. Design a pair of PCR primers that will amplify just the ORF in this sequence Design a pair PCR primers that will amplify just the ORF and add the following sequence to the 5' end of the amplified DNA fragment: GAATTC (what is this?) 3. Design a pair PCR primers that will amplify just the ORF and add the following sequence to the 3' end of the amplified DNA fragment: GGATCC (what is this?) 4. Design a pair PCR primers that will amplify the region from the start codon to the 5" cysteine 5. Design a pair PCR primers that will amplify the region from the 5" cysteine to the stop codon 6. Design a pair PCR primers that will amplify the region from the start codon to the 5 cysteine, but will change the cysteine codon to a glycine codon. 7. Design a pair PCR primers that will amplify the region from the 5 cysteine to the stop codon, but will change the cysteine codon to a glycine codon

Explanation / Answer

1.

Forward: 5'-atggctaaacatcat-3'

Reverse: 3'-aagttcttctccact-5'

2. GAATTC is the restriction site for EcoRI.

Forward: 5'-gaattcatggctaaa-3'

Reverse: 3'-aagttcttctccact-5'

3.

Forward: 5'-atggctaaacatcat-3'

Reverse: 3'-ttctccactcttaag-5'

4.

Forward: 5'-atggctaaacatcat-3'

Reverse: 3'-ttcacacactaaaca-5'

5.

Forward: 5'-tgtgactcctatgtg-3'

Reverse: 3'-aagttcttctccact-5'

6.

Forward: 5'-atggctaaacatcat-3'

Reverse: 3'-ttcacacactaaCCA-5'

7.

Forward: 5'-GGTgactcctatgtg-3'

Reverse: 3'-aagttcttctccact-5'

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