BackgroundColorectal cancer is the third most common type of cancer in men and women and the second leading cause of cancer-related deaths in the United States and UK. Colorectal cancer is strongly related to age, with almost three-quarters of cases occurring in people aged 65 or over. Pre-symptomatic screening is one of the most powerful tools for preventing colorectal cancer. Recently, the use of mitochondrial tRNA genes mutation or polymorphism patterns as a biomarker is rapidly expanding in different cancers because tRNA genes perform several functions including processing and translation which are essential components of mitochondrial protein synthesis. The aim of the present study was to find out the association of mitochondrial A12308G alteration in tRNALeu(CUN) in colorectal cancer and its usage as a new biomarker screening test.MethodsA tumor tissues from 30 patients who had colorectal cancer were selected randomly. The A12308G alteration in tRNALeu (CUN) was screened in the 30 colorectal tumor tissues. For comparison, 100 blood samples of healthy controls using PCR-sequencing methods were selected and the following results were found.ResultThe A12308G, a polymorphic mutation in V-loop tRNALeu(CUN), was found in 6 Colorectal tumor tissues and 3 healthy controls. A statistical significant difference was found between cases and control regarding the association of the A12308G mutation with the colorectal tumor (P < 0.05).ConclusionsThe A12308G, a polymorphic mutation in V-loop tRNALeu(CUN), could be considered as pathogenic mutation in combination with mitochondrial external conditions and other mitochondrial genes in developing different diseases especially cancers and could be used as one of the diagnostic tool. Also it seems that maybe there is relevance between A12308G mutation and other mutations that it can cause various phenotypes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13000-015-0337-6) contains supplementary material, which is available to authorized users.