Extensive scanning tunneling microscopy and spectroscopy experiments complemented by firstprinciples and parametrized tight binding calculations provide a clear answer to the existence, origin, and robustness of van Hove singularities (vHs) in twisted graphene layers. Our results are conclusive: vHs due to interlayer coupling are ubiquitously present in a broad range (from 1 to 10 ) of rotation angles in our graphene on 6H-SiC(000-1) samples. From the variation of the energy separation of the vHs with the rotation angle we are able to recover the Fermi velocity of a graphene monolayer as well as the strength of the interlayer interaction. The robustness of the vHs is assessed both by experiments, which show that they survive in the presence of a third graphene layer, and by calculations, which test the role of the periodic modulation and absolute value of the interlayer distance. Finally, we clarify the role of the layer topographic corrugation and of electronic effects in the apparent moiré contrast measured on the STM images. DOI: 10.1103/PhysRevLett.109.196802 PACS numbers: 73.22.Pr, 61.48.Gh, 68.37.Ef, 73.20.At Soon after the discovery of the unique electronic properties of graphene [1-3], suggestions were made for engineering the band structure of this material. It has been proposed that periodic potentials with wavelengths in the nanometer range could lead to anisotropic renormalization of the velocity of low energy charge carriers [4] or to the generation of new massless Dirac fermions [5]. Experimental works intended for verifying these theoretical predictions were recently reported [6][7][8], where the periodic perturbation was generated either by a lattice mismatch with the supporting material or by a self-organized array of clusters. An alternative route for modifying graphene's band structure would be to exploit a rotation between stacked graphene layers [9]. According to calculations, for large angles ( !15 ) the low energy band structure of graphene should be preserved [10][11][12]. For intermediate angles (1 15 ), it is predicted that, while the linear dispersion persists in the vicinity of the Dirac points of both layers, the band velocity is depressed and two saddle points appear in the band structure, giving rise to two logarithmic van Hove singularities (vHs) in the density of states (DOS) [9,[13][14][15][16][17][18]. For smaller angles ( 1 ) weakly dispersive bands appear at low energy [19,20] with sharp DOS peaks very close to the Dirac point [17,18].Twisted graphene layers are commonly found on different substrates, such as metals [13,21,22], the C face of SiC [23][24][25], or graphite surfaces [26,27]. Transfer techniques yielding large domains of twisted bilayers over a macroscopic sample [28] and quantitative, fast, Raman characterization tools [29,30] have recently been proposed. However, despite the fact that rotated graphene layers are readily available and a number of measurements have confirmed that large twist angles lead to an electronic decoupling of stacked graphene layers [...