To further analyze the CRABS CLAW gene, you create a map of the genomic clone. The 11-kb EcoRI fragment is ligated into the EcoRI site of the MCS of the vector shown in Problem 18. You digest the double-stranded form of the genome with several restriction enzymes and obtain the following results. Draw, as far as possible, a map of the genomic clone of CRABS CLAW.

What restriction digest would help resolve any ambiguity in the map?

Verified step by step guidance

1. Step 1: Begin by analyzing the data provided for each restriction enzyme digest. Note the fragment sizes generated by EcoRI, XbaI, XhoI, SalI, and HindIII individually, as well as the combinations of EcoRI with each enzyme. This will help identify overlapping fragments and potential cut sites.
2. Step 2: Construct a preliminary map of the genomic clone based on the fragment sizes. Start with the EcoRI digest, which produces fragments of 11.0 kb and 3.0 kb. Place these fragments on the map, ensuring their total length matches the genomic clone size.
3. Step 3: Incorporate the results of the double digests (e.g., EcoRI + XbaI, EcoRI + XhoI, etc.) into the map. For example, the EcoRI + XbaI digest produces fragments of 4.5 kb, 6.5 kb, and 3.0 kb. Use this information to determine the relative positions of XbaI cut sites within the EcoRI fragments.
4. Step 4: Repeat the process for the other double digests (EcoRI + XhoI, EcoRI + SalI, EcoRI + HindIII). Compare the fragment sizes generated by each combination to refine the map and resolve ambiguities in the placement of restriction sites.
5. Step 5: To resolve any remaining ambiguities, consider performing additional restriction digests with enzymes not yet used or combinations of enzymes that target specific regions of the map. For example, a digest with XbaI + XhoI or SalI + HindIII could provide further clarity on overlapping fragments or closely spaced cut sites.

Key Concepts

Restriction Enzymes

Restriction enzymes, or restriction endonucleases, are proteins that cut DNA at specific sequences, allowing scientists to manipulate and analyze genetic material. Each enzyme recognizes a unique sequence of nucleotides, which is crucial for creating DNA fragments of desired lengths for cloning or mapping. Understanding how these enzymes work is essential for interpreting the results of a restriction digest and constructing a genomic map.

Genomic Mapping

Genomic mapping involves determining the locations of genes and other important sequences on a chromosome. This process often utilizes restriction enzyme digests to create a physical map of DNA fragments, which can be analyzed to understand gene organization and function. A well-constructed map helps resolve ambiguities in gene locations and relationships, making it a fundamental aspect of genetic analysis.

Fragment Size Analysis

Fragment size analysis is a technique used to determine the lengths of DNA fragments generated by restriction enzyme digestion. By comparing the sizes of these fragments, researchers can infer the positions of restriction sites and the overall structure of the genomic clone. This analysis is critical for resolving ambiguities in genomic maps, as different combinations of restriction enzymes can yield distinct patterns that clarify the arrangement of genetic elements.