In which direction does DNA polymerase I primarily break down the RNA primer via exonuclease function?

• A. 3' → 5'
• B. Both directions
• C. It does not break down primers
• D. 5' → 3'

Answer: (D)

During lagging-strand synthesis, the RNA primer must be removed and replaced with DNA. DNA polymerase I has a 5'→3' exonuclease activity that chews away RNA nucleotides starting from the primer’s 5' end, while its 3'→5' exonuclease activity handles proofreading of DNA. This 5'→3' exonuclease action is what clears the RNA primer, creating a gap that the polymerase activity fills with DNA, after which ligase seals the nick. Therefore, the primer is removed in the 5' to 3' direction. The 3'→5' direction is not used for primer removal, and primer removal is not bidirectional.