What is the purpose of T4 DNA ligase in cloning, and what is meant by making cells competent for transformation?

What is the purpose of T4 DNA ligase in cloning, and what is meant by making cells competent for transformation?

• A. Ligase seals the DNA backbone; competent cells have permeable membranes allowing plasmid DNA uptake during transformation.
• B. Ligase unwinds DNA; competent cells replicate plasmids.
• C. Ligase degrades DNA; competent cells cannot take up DNA.
• D. Ligase synthesizes RNA; competent cells are specialized for transcription.

Answer: (A)

This item tests two ideas: how T4 DNA ligase functions in cloning and what it means for cells to be competent for transformation. In cloning, T4 DNA ligase seals the DNA backbone by forming phosphodiester bonds between adjacent DNA fragments, effectively joining a plasmid vector with an insert when their ends are compatible. This creates a covalently closed recombinant plasmid that can be replicated in bacteria. If ligase isn’t used, the fragments remain unconnected and cannot be stably inherited or expressed. Making cells competent means preparing them to take up DNA from the surrounding environment. Competent cells have membranes (or cell envelopes) made permeable to plasmid DNA, allowing transformation to occur. This competence is typically achieved chemically, often with calcium ions and a brief heat shock, or by electrical methods (electroporation) that temporarily permeabilize the cells. Once DNA enters, the plasmid can be expressed and propagated in the transformed cells.

This item tests two ideas: how T4 DNA ligase functions in cloning and what it means for cells to be competent for transformation. In cloning, T4 DNA ligase seals the DNA backbone by forming phosphodiester bonds between adjacent DNA fragments, effectively joining a plasmid vector with an insert when their ends are compatible. This creates a covalently closed recombinant plasmid that can be replicated in bacteria. If ligase isn’t used, the fragments remain unconnected and cannot be stably inherited or expressed.

Making cells competent means preparing them to take up DNA from the surrounding environment. Competent cells have membranes (or cell envelopes) made permeable to plasmid DNA, allowing transformation to occur. This competence is typically achieved chemically, often with calcium ions and a brief heat shock, or by electrical methods (electroporation) that temporarily permeabilize the cells. Once DNA enters, the plasmid can be expressed and propagated in the transformed cells.