In DNA data storage, the massive sequence complexity creates challenges in repeatable and efficient information readout. Here, our study clearly demonstrated that canonical polymerase chain reaction (PCR) created significant DNA amplification biases, which greatly hinder fast and stable data retrieving from hundred-thousand synthetic DNA sequences encoding over 2.85 megabyte (MB) digital data. To mitigate the amplification bias, we adapted an isothermal DNA amplification for low-bias amplification of DNA pool with massive sequence complexity, and named the new method isothermal DNA reading (iDR). By using iDR, we were able to robustly and repeatedly retrieve the data stored in DNA strands attached on magnetic beads (MB) with significantly decreased sequencing reads, compared with the PCR method. Therefore, we believe that the low-bias iDR method provides an ideal platform for robust DNA data storage, and fast and reliable data readout.