As a unicellular eukaryotic organism, Tetrahymena behaves a rapid and sensitive response to xenobiotics. A large number of gene families that are involved in some core processes associated with the sensing of and responding to environmental changes are presented within its genome. Thus, Tetrahymena can serve as an ideal model system to study the origin, evolutionary and functional diversity of eukaryotic gene families. In the current study, two well-conserved gene families — Metallothionein and Cytochrome P450, which are closely associated with the cellular physiological process under environmental stress, were investigated. Analyses were carried out on the aspects such as gene structure, gene function and evolutionary relationships by cloning and identification, expression characterization and evolutionary calculations: 1. A new multi-stress inducible metallothionein gene isoform (TpMT-2) was cloned and characterized from T. pyriformis. The putative protein sequence of TpMT-2 contains typical characteristics of the Tetrahymena Cd-inducible MT genes, but the sequence has a special feature of four intragenic tandem repeats within its first half. To investigate the transcriptional activities of both TpMT-2 and TpMT-1 genes toward heavy metals and H2O2, the mRNA levels of these two isoforms were evaluated by means of real-time quantitative PCR. Results showed that Cd was the most effective metal inducer of both two isoforms, but the relative expression level of TpMT-2 was much lower than that of TpMT-1. Zn was found to have the transcriptional inducing effect on both Tetrahymena MT genes. It was suggested that TpMT-1 and TpMT-2 have different physiological roles. The putative unique structural characterization and evolutionary mode of the MT 7a subfamily in the Tetrahymena genus are discussed; 2. 44 putative cytochrome P450 genes with full-length ORFs in the T. thermophila genome were identified by computer analyses coupled with cDNA sequence cloning methods. A good correlation between the conservation of intron position and phylogenetic relationships of T. thermophila P450 subfamily members was indicated; therefore, it can be inferred that the enlargement of several P450 families in T. thermophila resulted from recent separate small duplication events. The gene expression patterns during three important cell physiological cycles (vegetative growth, starvation and conjugation) were analyzed based on the EST and microarray data. For the T. thermophila P450 genes, there is no simple correlation between the expression level and the use of a preferred set of codons. The site-specific selection test showed that different rate of substitutions were estimated to occur within the coding regions among the CYP5005 and CYP5010 family members. We further discussed both the significance and limitation of evolutionary analysis for understanding the characteristics and functional diversities of the T. thermophila P450 isoforms.