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Early embryonic loss, occurring just prior to implantation is a significant cause of reproductive wastage in cattle. Dietary supplementation with n-3 polyunsaturated fatty acids (n-3 PUFA) has long been postulated to have a positive effect on fertility and reproductive success. The objective of the study was to examine (i) the effect of dietary supplementation with n-3 PUFA and level of post-insemination plane of nutrition on the uterine endometrial transcriptome and (ii) to identify alterations in key genes and pathways that may affect pregnancy outcome. A total of 60 estrous synchronized crossbred beef heifers were fed a high energy diet on an ad libitum basis and randomly assigned to one of two groups where the concentrate portion of the diet was either supplemented with a partially rumen protected source of n-3 PUFA (n = 32) or was unsupplemented (Control; n = 28) for 30 days prior to insemination. Immediately following insemination, animals were further allocated one of two post insemination diets, either remaining on the high plane of nutrition (High; n = 31) or were offered 60% of estimated maintenance energy requirements (Low; n = 29). Heifers were maintained on their respective diets until slaughter and embryo recovery on Day 16 post insemination or pregnancy diagnosis by ultrasonic scanning on Day 30. Uterine endometrial tissue was collected from slaughtered heifers at Day 16, RNA isolated, and gene expression analysis conducted by RNAseq. There was no effect of either supplementation with n-3 PUFA or post-insemination plane of nutrition on pregnancy rate at either Day 16 or 30. Comparison of transcript abundance across groups, however, highlighted a statistically significant effect of diet on uterine endometrial transcript levels, with a notable effect of n-3 PUFA supplementation on a number of differentially expressed genes (DEG). Of particular interest, the comparison of n-3 PUFA supplemented and unsupplemented pregnant heifers on the low plane of post-insemination nutrition resulted in 561 DEG, including the increased expression of genes previously demonstrated to be involved in early pregnancy, such as UPK3BL and CTSV, and in reproductively important pathways, including an embryonic development pathway and mTOR signaling pathway enriched in PUFA supplemented heifers, important to establishing pregnancy. Results indicate that despite no effect on pregnancy outcome, supplementation with n-3 PUFA positively altered a number of key fertility related genes and pathways involved in early pregnancy, which may have roles in the maintenance of pregnancy. [ABSTRACT FROM AUTHOR]