<p><i>Gibberella fujikuroi</i> QJGA4-1, a filamentous fungus, is recognized for its production of various gibberellic acids (GAs) secondary metabolites, such as GA₃, GA₄, GA₇, and GA₉. Among these, GA₄₊₇ has attracted considerable interest due to its distinct bioactivity. Its generally low yield level during fermentation presents a challenge for large-scale production. In our investigation, a combination of atmospheric and room temperature plasma (ARTP) mutagenesis and antibiotic stress screening was used to obtain a high-yielding mutant strain. Following a two-step mutagenesis, the GA₄₊₇ production of mutant strain <i>G. fujikuroi</i> M-1 reached 373.12 ± 14.23&#xa0;mg/L, marking a 14.41% increase compared to the wild-type (WT) strain. The result of scanning electron microscope indicated that the hyphae of <i>G. fujikuroi</i> M-1 were thinner than that of WT strain, suggesting that ARTP mutagenesis affected both cell growth and GA₄₊₇ biosynthesis of <i>G. fujikuroi</i>. In addition, one-factor-at-a-time experiments were carried out to initially assess carbon sources, nitrogen sources, and inorganic salts. This was succeeded by optimizing the fermentation medium by using response surface methodology, resulting in an elevated GA₄₊₇ production of 780.10 ± 30.00&#xa0;mg/L in <i>G. fujikuroi</i> M-1, which showed an 120.96% increase in production of GA₄₊₇ compared to that produced by <i>G. fujikuroi</i> QJGA4-1 in original culture medium. Combining ARTP mutation with medium optimization is an effective approach for improvement of GA₄₊₇ production.</p>