what could explain the migration of a protein in a gel at 100kD, when by DNA sequencing, the protein should be 60kD?
what could explain the migration of a protein in a gel at 100kD, when by DNA sequencing, the protein should be 60kD?
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You further investigate the regulatory proteins that bind to Regions 1, 2 and 3 using gel shift assays. A gel shift assay uses labeled DNA as a target for regulatory protein binding.
A labeled DNA fragment that fails to bind a protein will migrate quickly on an electrophoretic gel.
A labeled DNA fragment that does bind a protein will migrate more slowly because the DNA + protein complex is larger than the DNA alone. Thus, the label associated with the DNA will âshiftâ to a higher location on the gel, as shown in this diagram.
To investigate regulatory protein binding to Regions 1, 2, and 3, you will use labeled DNA fragments with some of the same point mutations as in Part B, incubate them with proteins extracted from heart or lung tissue, and conduct a gel shift assay. How would gel shift assay results be affected by these mutations?
Select the two correct answers.
| 1. Region 2 binds protein from lung tissue. If Region 2 is mutated, then the DNA will not shift on the gel when incubated with protein from lung tissue. |
| 2. Regions 1 and 3 bind protein from heart tissue. If both Regions 1 and 3 are mutated, then the DNA will not shift on the gel when incubated with protein from heart tissue. |
| 3. Region 2 binds protein from heart tissue. If Region 2 is mutated, then the DNA will not shift on the gel when incubated with protein from heart tissue. |
| 4. Regions 1 and 3 bind protein from lung tissue. If both Regions 1 and 3 are mutated, then the DNA will not shift on the gel when incubated with protein from lung tissue. |
