I need an explanation of the following paragraph about LIC from a protein expression review article.
"For T4 DNA polymerase-based LIC, the 5’ extensions on each primer (of length 12-15 bases) are designed so that the sequence is composed of only three nucleotides, lacking the fourth. For example, when using the plasmid pNIC28-Bsa4 (SGC, GenBank accession EF198106), the following extensions are used in the primers: Upstream primer: TACTTCCAATCCATG (the ATG encodes methionine in-frame) Downstream primer: TATCCACCTTTACTGTCA (the TCA is a stop codon in reverse). In both cases, treating the purified PCR fragments with T4 DNA polymerase in presence of dCTP will result in 3’-5’ exonuclease activity, which will degrade the opposing strand down to the first C, which is opposite the first G (bold) in the primer sequence. The resulting single-stranded extensions precisely match such single stranded extensions in the vector. The 3’-exonuclease activity of T4 DNA polymerase is thus used to expose complementary single-stranded stretches on the vector and inserts. The treated vector and inserts are annealed and introduced directly into bacteria."
1- The 5' extensions added to the forward and reverse primers are supposed to match the vector sequence, right? My question is how it is even possible to create these extensions to be made of only 3 nucleotide types while these 5' extensions are supposed to match the vector sequence (most likely will have all 4 nucleotides)?
2- What is role played by methionine codon and the stop codon in the upstream and downstream primers respectively?
3-Why would the addition of dCTP activate the 3' endonuclease activity of T4 DNA polymerase?