Which polymerase is generally used in Sanger sequencing?
Classical Sanger sequencing traditionally used the Klenow fragment of Escherichia coli DNA polymerase I, which retains DNA polymerase and 3' to 5' proofreading exonuclease but lacks 5' to 3' exonuclease activity that would degrade primer. This made it ideal for controlled extension and chain termination without unwanted digestion. Later improvements adopted Sequenase, a chemically modified T7 DNA polymerase, and thermostable Taq variants for automated cycle sequencing. DNA ligase seals nicks rather than polymerizing, reverse transcriptase copies RNA. Klenow represents historically taught enzyme for this method.
Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.