You have isolated a genomic clone with an EcoRI fragment of 11 kb that encompasses the CRABS CLAW gene (see Problem 18). You digest the genomic clone with HindIII and note that the 11-kb EcoRI fragment is split into three fragments of 9 kb, 1.5 kb, and 0.5 kb.

Restriction enzyme sites within a cDNA clone are often also found in the genomic sequence. Can you think of a reason why occasionally this is not the case? What about the converse: Are restriction enzyme sites in a genomic clone always in a cDNA clone of the same gene?

Key Concepts

Restriction Enzymes

Restriction enzymes are proteins that cut DNA at specific sequences, known as recognition sites. They are essential tools in molecular biology for cloning and analyzing DNA fragments. Different enzymes recognize different sequences, which can lead to varying fragment sizes when digesting DNA, as seen in the question with EcoRI and HindIII.

Genomic vs. cDNA Clones

Genomic clones contain the entire genomic DNA, including introns and regulatory regions, while cDNA clones are synthesized from mRNA and represent only the expressed genes, excluding introns. This difference can lead to the absence of certain restriction sites in cDNA clones that are present in genomic DNA, particularly if the sites are located within introns.

Alternative Splicing

Alternative splicing is a process by which a single gene can produce multiple mRNA variants by including or excluding certain exons during transcription. This can result in different protein isoforms and may affect the presence of restriction enzyme sites in cDNA clones, as some sites may be located in exons that are not included in all mRNA variants.