Abstract The present study was conducted to investigate the gold bioleaching factors from ore by cyanide producing bacterium Chromobacterium violaceum. The optimal condition for cyanide production by C. violaceum was pH 9 and 5g/L of glycine in YP medium in 2-days of incubation. In shake flask culture, gold bioleaching from the ores by C. violaceum was determined with various experimental protocols such as particle size, pre-grown C. violaceum, pH, and biooxidative treatment. The three types of low grade ores viz., R, S, and H were used. The gold bioleaching efficiencies were recorded as 0%, 50%, and 5% for ores R, S, and H, respectively, when C. violaceum culture was used without any pretreatment (protocol 1). In the experimental protocol involving grinding and pre-grown C. violaceum, leaching efficiencies increased to 60%, 100%, and 40% for ore R, S, and H samples, respectively. Especially, the bioleaching efficiency of ore S enhanced to almost 100% with pre-grown C. violaceum (protocol 2) and that of ore R increased to 53% (i.e., 96% of cyanidable gold) with grinding (protocol 3) due to their mineralogical characteristics of the ores. For refractory gold (i.e., ore R) grinding as pretreatment was needed, and for ore S (almost all of the gold was cyanidable) cyanide production was activated by using pre-grown bacteria. Biooxidation with Acidithiobacillus ferrooxidans and pH adjustment (i.e., 9–11) did not affect the bioleaching efficiencies. The mineralogical cause of gold refractoriness was analyzed by automated SEM that showed most of gold in the ore was entrapped in pyrite and silica. The results indicated that gold bioleaching by C. violaceum from low grade ore can be enhanced by grinding and pre-grown microbe; use of appropriate experimental condition is important according to the mineralogical characteristics.