Multiple Choice
Each restriction enzyme has a specific sequence of nucleotides where it cuts the DNA. These sequences of DNA are unique to each restriction enzyme and are known as:
18. Biotechnology
Steps to DNA Cloning
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What is the most logical sequence of steps for splicing foreign DNA into a plasmid and inserting the plasmid into a bacterium?
I. Transform bacteria with a recombinant DNA molecule.
II. Cut the plasmid DNA using restriction enzymes (endonucleases).
III. Extract plasmid DNA from bacterial cells.
IV. Hydrogen-bond the plasmid DNA to non-plasmid DNA fragments. V. Use ligase to seal plasmid DNA to non-plasmid DNA.
A
II, III, V, IV, I
B
III, II, IV, V, I
C
III, IV, V, I, II
D
IV, V, I, II, III
Verified step by step guidance1
Step 1: Begin by extracting plasmid DNA from bacterial cells (Step III). This involves isolating the plasmid DNA, which will serve as the vector for inserting foreign DNA.
Step 2: Use restriction enzymes (endonucleases) to cut the plasmid DNA at specific recognition sites (Step II). This creates openings in the plasmid where foreign DNA can be inserted.
Step 3: Allow the plasmid DNA to hydrogen-bond with non-plasmid DNA fragments (Step IV). This step involves complementary base pairing between the sticky ends of the plasmid and the foreign DNA.
Step 4: Use DNA ligase to seal the plasmid DNA to the non-plasmid DNA fragments (Step V). Ligase catalyzes the formation of phosphodiester bonds, creating a stable recombinant DNA molecule.
Step 5: Transform bacteria with the recombinant DNA molecule (Step I). This involves introducing the recombinant plasmid into bacterial cells, where it can replicate and express the foreign DNA.
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