Does maternal ageing and ovarian stimulation alter mitochondrial DNA (mtDNA) copy number and gene expression of oocytes and cumulus cells from a novel bovine model for human IVF? Oocytes collected from females with identical nuclear genetics show decreased mtDNA copy number and increased expression of an endoplasmic reticulum (ER) stress gene with repect to ovarian stimulation, whilst differences in the expression of genes involved in mitochondrial function, antioxidant protection and apoptosis were evident in relation to maternal ageing and the degree of ovarian stimulation in cumulus cells. Oocyte quality declines with advancing maternal age; however, the underlying mechanism, as well as the effects of ovarian stimulation are poorly understood. Human studies investigating these effects are often limited by differences in age and ovarian stimulation regimens within a patient cohort, as well as genetic and environmental variability. A novel bovine cross-sectional maternal age model for human IVF was undertaken. Follicles were aspirated from young (3 years of age; n = 7 females) and old (10 years of age; n = 5 females) Holstein Freisian clones following multiple unstimulated, mild and standard ovarian stimulation cycles. These bovine cloned females were generated by the process of somatic cell nuclear transfer (SCNT) from the same founder and represent a homogeneous population with reduced genetic and environmental variability. Maternal age and ovarian stimulation effects were investigated in relation to mtDNA copy number, and the expression of 19 genes involved in mitochondrial function, antioxidant protection, oocyte-cumulus cell signalling and follicle development in both oocytes and cumulus cells. Young (3 years of age; n = 7 females) and old (10 years of age; n = 5 females) Holstein Freisian bovine clones were maintained as one herd. Stimulation cycles were based on the long GnRH agonist down-regulation regimen used in human fertility clinics. Follicle growth rates, numbers and diameters were monitored by ultrasonography and aspirated when the lead follicles were >14 mm in diameter. Follicle characteristics were analysed using a mixed model procedure. Quantitative PCR (qPCR) was used to determine mtDNA copy number and reverse transcriptase-qPCR (RT-qPCR) was used to measure gene expression in oocytes and cumulus cells. Method of ovarian stimulation (P = 0.04), but not maternal age (P > 0.1), was associated with a lower mtDNA copy number in oocytes. Neither factor affected mtDNA copy number in cumulus cells. In oocytes, maternal age had no effect on gene expression; however, ovarian stimulation in older females increased the expression of GRP78 (P = 0.02), a gene involved in ER stress. In cumulus cells, increasing maternal age was associated with the higher expression of genes involved in mitochondrial maintenance (TXN2 P = 0.008 and TFAM P = 0.03), whereas ovarian stimulation decreased the expression of genes involved in mitochondrial oxidative stress and apoptosis (TXN2 P = 0.002, PRDX3 P = 0.03 and BAX P = 0.03). The low number of oocyte and cumulus cell samples collected from the unstimulated cycles limited the analysis. Fertilization and developmental potential of the oocytes was not assessed because these were used for mtDNA and gene expression quantification. Delineation of the independent effects of maternal age and ovarian stimulation regimen on mtDNA copy number gene expression in oocytes and cumulus cells was enabled by the removal of genetic and environmental variability in this bovine model for human IVF. Therefore, these extend upon previous knowledge and findings provide relevant insights that are applicable for improving human ovarian stimulation regimens. Funding was provided by Fertility Associates and the University of Auckland. J.C.P. is a shareholder of Fertility Associates and M.P.G. received a fellowship from Fertility Associates. The other authors of this manuscript declare no conflict of interest that could be perceived as prejudicing the impartiality of the reported research.