We analyzed δ²⁹Si of dissolved silicate for eight water column profiles across the Southern Ocean (south of Australia in spring 2001) from the Seasonal Ice Zone (SIZ) north to the Subantarctic Zone (SAZ), including the first isotopic compositions measured for Si-depleted seawaters. All profiles display mixed layer enrichments in heavy Si isotopes relative to deep water in accordance with preferential uptake of the light isotope by diatoms. As silicate levels decrease from the SIZ northward across the Polar Front Zone (PFZ) to the SAZ, surface and mesopelagic δ²⁹Si signatures generally become progressively heavier, but the most Si-depleted SAZ waters do not exhibit δ²⁹Si values heavier than in the PFZ. This intricacy appears to derive from variations in the vertical and horizontal supply of silicate to surface waters, and by applying a steady state open system model, we estimate a fractionation factor, ²⁹ , between diatoms and seawater of −0.45 ± 0.17‰, independently of zones and phytoplankton community. Though encouraging, these results are related to latitudinal changes in mesopelagic δ²⁹Si values, complexity in surface silicate−δ²⁹Si correlations, and differences from previous studies, which underline the need for caution in the use of silicon isotopes in paleoceanographic studies until systematic efforts have been undertaken to better understand modern variations.