<![CDATA[Species identification within the family Felidae is often challenged by overlapping morphological characteristics resulting from hybridization, necessitating the use of genetic approaches. This study aimed to analyze the phylogenetic relationships and evolutionary history of 40 Felidae species based on mitochondrial Cytochrome b (CytB) gene sequences. This in silico study utilized secondary data obtained from GenBank, which were analyzed using MEGA 12 for sequence alignment, nucleotide composition analysis, and phylogenetic tree reconstruction using the Maximum Likelihood method with 1,000 bootstrap replicates, followed by validation using DrPhylo. The results showed a predominance of transition mutations, with a high R ratio (9), as well as significant codon usage bias. Phylogenetic reconstruction divided the family Felidae into eight major clades supported by high bootstrap values, namely Panthera, Neofelis, Pardofelis, Prionailurus, Lynx, Leopardus, Felis, and a combined Puma group. The genus Otocolobus was identified as the oldest lineage, diverging earliest from the other lineages. The M-Grid test confirmed the validity of the major clade groupings, with confidence levels approaching 1.00. It can be concluded that the CytB gene is an effective and reliable genetic marker for phylogenetic studies of the family Felidae.]]>
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