π-Conjugation plays an important role in charge transport through single molecular junctions. We describe in this paper the construction of a mechanically controlled break-junction setup (MCBJ) equipped with a highly sensitive log I–V converter in order to measure ultralow conductances of molecular rods trapped between two gold leads. The current resolution of the setup reaches down to 10 fA. We report single-molecule conductance measurements of an anthracene-based linearly conjugated molecule (AC), of an anthraquinone-based cross-conjugated molecule (AQ), and of a dihydroanthracene-based molecule (AH) with a broken conjugation. The quantitative analysis of complementary current–distance and current–voltage measurements revealed details of the influence of π-conjugation on the single-molecule conductance.