The middle and lower Yellow River regions were crucial for early agricultural development and the emergence of complex societies in East Asia, yet their fine-scale population history from the Neolithic to historical periods remains incompletely resolved. Here, we present genome-wide data from 61 ancient individuals (∼4,500–200 before present [BP]) from Henan and Shandong. Our analyses show that middle-to-late Neolithic populations at Dahecun in Henan are genetically homogeneous and closely related to contemporaneous Yellow River millet-farming populations. By ∼3,000 BP, Shandong populations shift toward a late Neolithic middle Yellow River-related ancestry profile, consistent with the eastward expansion of middle Yellow River agricultural populations. This ancestry profile persists into historical periods, indicating long-term genetic continuity in Shandong. Two Tang-Song-period individuals from western Shandong carry ∼15% West Eurasian-related ancestry, indicating limited cross-regional genetic interaction. These findings refine the population history of the middle and lower Yellow River regions.