Ligation-Independent Cloning (LIC) Methodology. Shows the general procedure for LIC (1) The primer sequences required for amplification of the gene of interest. (2) Both the vector and insert are subject to treatment with T4 DNA polymerase, in the presence of either dTTP (vector) or dATP (insert). (3) The result after successful annealing. (4) After cleavage with TEV protease, the tags are removed and the protein retains 4 amino acids (GAAS).