A company has created 3 types of tomatoes: red, pink and white. You want to unde

Question

A company has created 3 types of tomatoes: red, pink and white. You want to understand why these tomatoes have different colors. You know that the "red" gene contains a SNP in its promoter: The promoter sequence is shown below; the 33^rd nucleotide can be an A or a C. Because this SNP is in the TATA box of the red gene, you think that depending on whether this nucleotide is an A or a C, the gene will either be expressed (on = red allele) or remain silent (off= white allele). Based on this you generate a hypothesis that can possibly explain why tomatoes have different colors: if the red gene is off, the tomato will be white, if it is on it will be red and if only one allele is on, it will be pink. So you collect red, pink and white tomatoes and do several experiments to see if your hypothesis is true. The restriction endonucleases shown above recognize the following sequences: SspI: 5'-AATATT-3' CviQI: 5'-GTAC-3' BspCNI: 5'-CTGAG-3' Tsp509I: 5'-AATT-3' ApoI: 5'-AAATTT-3' MnlI: 5'-GAGG-3' Experiment 1: You want to determine if the presence of the SNP at nt.#33 affects tomato color: to do this experiment, you obtain genomic DNA from 10 tomatoes that are red, pink and white tomatoes. You amplify the promoter region shown in the previous page for each tomato by PCR. To determine the presence of the SNP by PCR-RFLP you choose a restriction endonuclease from the list shown in the previous page, and perform PCR-RFLP. Your results are shown below. Experiment la: Then you perform a second experiment. This time you use a different restriction endonuclease. Your PCR-RFLP results are shown below: Experiment lb: Assuming that your hypothesis is correct; based on these results, which endonuclease (among those shown in the previous page) could have been used in experiment lb? Please explain your logic briefly.

Explanation / Answer

The endonuclease used was most likely MnII as this is the only enzyme which has a restriction site downstream to the 33rd nucleotide where the SNP has taken place. Hence this may give rise to a different pattern.

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