The study of the transition of mesophase to ⊍-crystal (meso-⊍ transition) is of great significance for understanding the crystallization, morphology as well as processability of polypropylene materials. The structural evolution of mesomorphic isotactic propylene-1-butene (iPPBu) random copolymer during the continuous heating process was investigated using in situ Fourier transform infrared spectroscopy, differential scanning calorimetry, wide-angle X-ray diffraction and dynamic mechanical analysis. The presence of the butene comonomer influences the meso-⊍ transition behavior. Compared with the mesomorphic iPP homopolymer, the presence of butene comonomer reduces both onset temperature (T onset ) and end temperature (T end ) of the meso-⊍ transition, and narrows the transition temperature range. The content of the 3 1 helical sequence with 12 monomers remains invariant during the meso-⊍ transition, differing from that of mesomorphic iPP homopolymer. Similar to mesomorphic iPP homopolymer, the 3 1 helical sequence with monomer number n ≤ 12 is found to exist in the rigid amorphous fraction of iPPBu, but is different from that of a mesomorphic propylene-ethylene random copolymer. In the meso-⊍ transition (from 60 to 100 °C), the redshift of the peak position of the band at ca 841 cm −1 provides new information for the approaching of adjacent stems at the molecular level. Moreover, a new peak at around T end is found in the tan(⊐)-T plot, and is correlated with the meso-⊍ transition. The activation energy of the meso-⊍ transition of mesomorphic iPPBu is 32.5 kJ mol −1 , which is slightly lower than that of mesomorphic iPP homopolymer. This work sheds new light on further understanding of the meso-⊍ transition mechanism of iPPBu copolymer at the molecular level and elucidates the effect of butene comonomer on the meso-⊍ transition.