Grafting experiments were performed using Gossypium hirsutumaccession TM‑1 andG. barbadense accession Pima 3-79 to identify grafting and rootstock effects on boll weight (BW), seed numbers per boll (SNB), seed weight (SW), cottonseed crude oil (CO), myristic acid (MA), palmitic acid (PA), stearic acid (SA), palmitoleic acid (POA), heptadecanoic acid (HAD), oleic acid (OA), linolenic acid (LA), gamma-linolenic acid (GLA), alfa-linolenic acid (ALA), cis‑8,11,14-eicosatrienoic acid (CEA), cis‑4,7,10,13,16,19-docosahexaenoic acid (CDA), lignoceric acid (LCA), and nervonic acid (NA). The results showed that BW and SW could be improved in G. hirsutumscions. On the other hand, BW, SNB, and SW traits could be improved in G. barbadensescions. The CO of two tetraploid species is mainly comprised of PA, OA, and LA. Grafting and rootstock effects altered the levels of CO, PA, OA, LA, MA, ALA, and NA. The results revealed that G. barbadensecould be used as the rootstock in homografts and heterografts. To figure out the molecular mechanism(s) behind fatty acid changes in grafts, more studies with cotton genotypes with different amounts of fatty acids are needed.