It has been recently demonstrated that free DNA tracers have the potential in tracing water flow and contaminant transport through the vadose zone. However, whether the free DNA tracer can be used in flood irrigation area to track water flow and solute/contaminant transport is still unclear. To reveal the infiltration process and retention mechanisms of surface applied free DNA tracer through soil under flood irrigation, we tested the fate and transport behavior of surface applied free DNA tracers through packed saturated sandy soil columns with a 10 cm water head mimicking flood irrigation. From the experimental breakthrough curves and by fitting a two-site kinetic sorption model (R² = 0.83-0.91 and NSE = 0.79-0.89), adsorption/desorption rates could be obtained and tracer retention profiles could be simulated. Together these results revealed that 1) the adsorption of free DNA was dominantly to clay particles in the soil, which took up 1.96 % by volume, but took up >97.5 % by surface area and densely cover the surface of sand particles; and 2) at a pore water pH of 8.0, excluding the 4.9 % passing through and 3.1 % degradation amount, the main retention mechanisms in the experimental soil were ligand exchange (42.0 %), Van der Waals interactions (mainly hydrogen bonds), electrostatic forces and straining (together 44.7 %), and cation bridge (5.3 %). To our knowledge, this study is the first to quantify the contribution of each of the main retention mechanisms of free synthetic DNA tracers passing through soil. Our findings could facilitate the application of free DNA tracer to trace vadose zone water flow and solute/contaminant transport under flood irrigation and other infiltration conditions.