## 3 Discussion  
The \(CYP\) gene family has been reported in numerous studies of some plant species with genome-wide characterization [7; 37]. The number of \(CYP\) genes ranges from 79 in cotton to 355 in durian (Table S11). In the present study, 276 \(O\)/\(CY\)Ps were identified from the \(O\). _fragrans_ reference genome and clustered into seven clans, which were further categorized into 40 families (Figure S1A). There were fewer \(CYP\) genes in \(O\). _fragrans_ than in most woody plants. Gene-duplication events, especially tandem duplication, are considered to be one of the main reasons for gene family expansion during the process of evolution [38]. A total of 67 tandem duplicated genes and 66 segmental duplication pairs with 163 \(O\)/\(CY\)Ps were  
Figure 9: Functional characterization of \(O\)/\(CYP142\) by stable transformation. (**A**) Semiquantitative RT-PCR to determine the expression intensity of \(O\)/\(CYP142\); a fragment of the \(NEE\)-\(1\alpha\) gene was amplified as the normalizer, \(\mathrm{EVI}\)–3 denote different plant lines that were stably transformed with empty vectors, and \(\mathrm{L1}\)–\(\mathrm{l2}\) denote different plant lines that were stably transformed with 355:O/\(CYP142\):GUS vectors. (**B**) Score scatter plot of the flower GC–MS profiles of two types of flowers with overexpression of \(O\)/\(CYP142\) and empty vector in tobacco, determined with PCA-X analysis in SIMCA14.1. Each type of plants consisted of three replicates. (**C**) Score scatter plot of the flower GC–MS profiles of two types of flowers with overexpression of \(O\)/\(CYP142\) and empty vector in tobacco, determined with OPLS-DA analysis in SIMCA14.1. (**D**) Loading scatter plot analysis of floral volatile organic compounds in plants that were stably transformed with 355:O/\(CYP142\):GUS and empty vectors. (**E**) Floral composition analysis of linealod after stable transformation with 355:O/\(CYP142\):GUS and empty vectors. (**F**) Floral composition analysis of leaf alcohol after stable transformation with 355:O/\(CYP142\):GUS and empty vectors. (**G**) Floral composition analysis of (-)-caryophyllene oxide after stable transformation with 355:O/\(CYP142\):GUS and empty vectors. (**H**) Information of key aromatic components selected based on VIP from OPLS-DA (VIP \(>\) 1);* significant difference at \(p<\) 0.05 and ** significant difference at \(p<\) 0.01. The lowercase letters above the columns represent the significant differences as assessed by [PERSON]’s test at the 0.05 level. identified in our study (Figure 3), indicating that evolution of the _OfCYP_ family may also have been driven by gene-duplication events. The conserved heme-binding domain (FoxGxRxCxG) and the PERF consensus (PXRX) are P450 signature motifs [39]. Motifs 1 and 3, representing the heme-binding domain and the PERF consensus, respectively, were present in every OfCYP protein. Most OfCYP proteins within the same can exhibited similar motif components but with significant differences among clans (Figure S1 and Table S2), indicating that the OfCYP members within the same family may have similar functions, and that some motifs may play a vital role in family-specific functions. The similar motif distribution patterns within the same family also imply that the identification and division of _OfCYP_ gene families in this study is reliable. Previous studies have shown that _CYP_ genes are widely distributed and participate in the synthesis of many metabolites, especially secondary metabolites [40]. It is well known that genes expressed specifically in certain tissues can have tissue-specific functions. The analysis of RPKM values from the transcriptome data from seven different organs provided organ-specific expression data for the _OfCYP_s (Figure S2 and Table S4), implying that these genes may play different roles in various organs in _O. fragrans_. _AtCYP73A5_ which plays an essential role in phenylpropanol metabolism, was expressed more in flowers than other organs [41].