**3. Results**

*3.1. Identification, Physiochemical Properties, Chromosomal Distribution, Phylogenetic Analysis, and Subcellular Localization*

In total, seventeen (17) SAMS genes were identified in the *Gossypium hirsutum* species using the NAU assembly. They are distributed on all of the fourteen (14) chromosomes of the *G. hirsutum* genome, with the chromosomes At11 and Dt11 harboring, respectively, two *GhSAMSs* (Figure 1a). One of them is located on a scaffold. We also investigated the chromosomal distribution of *GhSAMSs* in the genomes of *G. arboretum* (A genome) and *G. raimondii* (D genome). We found that the *GhSAMS* genes are located on chromosomes At02, At04, At07, At08, At09, At11, and At12 of the A genome and on chromosomes Dt02, Dt04, Dt07, Dt08, Dt09, Dt11, and Dt12 of the D genome (Figure 1b,c).

The physicochemical properties of the *GhSAMS* genes are presented in Table 2. Their protein sequence length ranged from 256 (*Gh\_A07G1193*) to 393 aa (*Gh\_A08G1067*), with a molecular weight (MW) varying from 28.12 to 42.61 kDa. Both the proteins were stable and exhibited negative GRAVY values. The isoelectric points (*PI*) of the protein ranged from 5.579 to 8.974 (Table 2).

The phylogenetic analysis of *GhSAMS* proteins and SAMS proteins from other related species was performed to examine their relationships (Figure 1d). The results indicated that SAMS proteins could be classified into five major groups (I–V). The Group III genes were all from monocotyledonous plant species, while the genes from the dicotyledonous plant species were scattered across all four groups. *GhSAMS* genes were classified into groups I, II, and IV, where they showed a high degree of similarity with the SAMS genes from soybean, *T. cacao*, and *M. truncatula* (Figure 1d). The subcellular localization analysis showed that SAMS proteins are located mainly in the cytoplasm and cytoskeleton of *G. hirsutum* cells (Table 2; Figure 1e,f).


**Table 2.** The physicochemical properties of cotton SAMS genes.

**Figure 1.** Chromosomal distribution, phylogenetic analysis, and subcellular localization of *GhSAMS* genes. (**a**–**c**) Distribution of *GhSAMS* genes on the chromosomes of *G. hirsutum, G. arboreum*, and *G. raimondii*, respectively. (**d**) Phylogenetic tree of *GhSAMS* genes and their homologs from *A. thaliana*, *T. cacao*, Soya bean, Rice, Tomato, *Medicago truncatula*, Sorghum, Maize, and Barley. (**e**,**f**) Subcellular localization of *GhSAMS* genes. The neighbor-joining method was used to construct the phylogenetic tree with replicates of 1000 bootstrap values in the MEGA 7.0 software.
