With reference to the map of pAP3neo given in Figure 2.2, and using the sequence given in section B, what are the expected sizes of a double digest using NruI and MluI restriction enzymes?

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Question restatement: The prompt asks, with reference to the map of pAP3neo in Figure 2.2 and using the sequence described in section B, what are the expected sizes of a double digest using NruI and MluI restriction enzymes?

Option A: 140 and 5210 bp fragments
- This pairing does not match the provided answer set and would imply a discrepancy with the map or the sequence information in section B. The sizes 140 bp and 5210 bp would require specific cut positions that may not align with the NruI and MluI sites in the described map. If the map indicates different fragment lengths, this option would be inconsistent.

Option B: 152 and 5198 bp fragments
- This option corresponds to the fragment sizes given in the provided answer field. If the map in Figure 2.2 shows NruI and MluI cut sites that generate a ~152 bp fragment and a ~5198 bp fragment when the double digest is performed, this would be consistent with the described map and the sequence in section B. The accuracy depends on the precise locations of the restriction sites on pAP3neo as depicted.

Option C: not enough information to answer
- This choice asserts that the available map and sequence do not provide sufficient data to determine fragment sizes. If Figure 2.2 explicitly presents the map with the locations of NruI and MluI sites and section B provides the sequence context needed to calculate fragment lengths, then there is enough information. However, if any essential details are missing (e.g., exact cut positions or a clearly annotated map), this would be a valid concern. The validity of this option hinges on whether the necessary coordinates are indeed present.

Option D: 112 and 5238 bp fragments
- This option proposes a different pair of fragment lengths. To evaluate it, one would need to verify whether the NruI and MluI sites fall at positions that yield ~112 bp and ~5238 bp fragments in the double digest. If the map indicates these specific cut sites, this could be plausible; otherwise, it would be inconsistent with the depicted restriction pattern.

Overall item-by-item consideration: The question relies on interpreting a labeled map (Figure 2.2) and sequence details (section B). The correctness of any option depends on whether the reported fragment sizes match the actual distances between NruI and MluI cut sites on pAP3neo as shown. If the figure and section provide exact coordinates or a calculated digest, then the option that aligns with those data is the valid one. If any of the data are ambiguous or missing, Option C would be the safest choice.

Notes on potential misconceptions: Some students might assume any small fragment (e.g., around 100–150 bp) is plausible for a double digest, but fragment sizes must correspond to the precise distance between adjacent restriction sites on the plasmid. Others might rely on approximate sizes without checking the map—this can lead to selecting an option that seems reasonable but is not consistent with the annotated sites. Finally, confusing NruI and MluI site locations can lead to incorrect pairings of fragment lengths; always verify the actual positions on the map and the sequence context from section B.

BCMB20005_2025_SM1 Week-3 Pre-Lab Quiz

With reference to the map of pAP3neo given in Figure 2.2, and using the sequence given in section B, what are the expected sizes of a double digest using NruI and MluI restriction enzymes?

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