A series of novel donor-acceptor conjugated alternating copolymers based on the isoindigo acceptor moiety have been designed, synthesized, and characterized, in order to explore the potential of isoindigo for efficient donor materials with high photovoltages in solar cells. We have systematically investigated and discussed the effect of combining different electron-rich thiophene-based units on the structural, optical, electronic, and photovoltaic properties of the resulting polymers. Morphological studies and quantum-chemical calculations are carried out to gain insights into the different properties. The power conversion efficiencies (PCEs) of the solar cells based on these polymers are increased step by step by over 3-fold through a rational structural modification. Among them, PBDTA-MIM shows a PCE of 5.4%, which is to our knowledge the best result achieved among isoindigo-based polymers for solar cells combined with PC61BM as the acceptor using the conventional device configuration. Our results further emphasize the use of isoindigo as an effective acceptor unit and highlight the importance of carefully choosing appropriate chemical structure to design efficient donor-acceptor polymers for organic solar cells. In addition, the resulting low optical gaps, the promising PCEs with PC61BM as the acceptor, and the good open-circuit voltages (up to 0.8 V) synergistically demonstrate the potential of this class of polymers as donor materials for bottom subcells in organic tandem solar cells.