Abstract: Objective Analyzing the mtDNA copy number in gastric cancer tissues and adjacent non-cancerous tissues, to investigate the relationship betweenmtDNA alteration and the clinicopathological parameters and patient prognosis in gastric cancer.Methods A total of 123 gastric cancer tissues and adjacent non-cancerous tissues were obtained from the tumor bank of Zhongnan Hospital of Wuhan Universitybetween 2011 and 2013. Relative mtDNA copy number was measured in triplicate by a quantitative real-time PCR assay. Using the statistic analysis analyzed the relationship between mtDNA content and clinicopathological features of the gastric cancer. Results The data showed that the majority of the cancer patients had low levels of mtDNA copy number as compared to adjacent non-cancerous tissues (P<0.0001). While there was no association of mtDNA content with the clinicopathological features, such as gender, age, tumor localization, differentiation, tumor invasion, lymph node metastasis,clinical stage, Her-2 expression, Ki-67 status. Additionally, by using the median ratio copy number as the cutoff point, the patients were separated into high and lowcopy number tumor groups, we found thatcompared with high mtDNA content, low mtDNA content was associated with T1+T2 group (P=0.04) and Ki-67negative (P=0.015). Conclusions In this study, the relative mean mtDNA content was lower in gastric cancer tissues than theadjacent non-cancerous tissues. Additionally, we found a strong link between increased mtDNA content and invasion of the tumor and Ki-67 status.We hypothesis that mtDNA copy number can be used as a new biomarker,and increasing the relative mtDNAcontent seems to be linked to the advancement of gastric cancer progression.