Milk fat (MF) triacylglycerol composition varies within a population of dairy cows. The variability of MF triacylglycerols and their structure was partially explained by the fatty acid (FA) composition of the MF, and by DGAT1 K232A polymorphism. The FA C16:0 and C18:1cis-9 play a major role in understanding the changes seen in triacylglycerol profile and structure because they are the most abundant FAs in MF and are negatively correlated. MFs with low ratio C16:0/C18:1cis-9 were decreased in triacylglycerols with 34 and 36 carbons and were increased in triacylglycerols with 52 and 54 carbons. These changes in MF composition greatly affected the crystallization behavior of MF by changing the types of polymorphs formed during its crystallization. MF with low ratio C16:0/C18:1cis-9 formed stable and metastable polymorphs (β and β’, respectively), whereas MF with high ratio C16:0/C18:1cis-9 formed exclusively metastable polymorphs (β’) when the fat was crystallized at 20°C. The changes in MF composition also affected the melting behavior of MF by changing the melting point of the MF fractions.