# Functional Analysis of Two Terpene Synthase Genes Isolated from the Flowers of _Hosta_ 'So Sweet'
## 3 Results
### Terpenoids Production in Hosta and Expression Analysis of HsTPS1 and HsTPS2 Genes  
Terpenoids are the dominant scent components in _Hosta_ flowers, mainly including linalool, myrcene, limonene, and (_E_)-\(\beta\)-ocimene [21]. The volatile compounds in flowers of \(H\). 'So Sweet' were analyzed by hexane extraction and GC-MS. The test results showed that the relative content of terpenoids in \(H\). 'So Sweet' was 91.78-93% of the total volatile compounds, with a small amount of phenylpropionic acid compounds and fatty family compounds. Fifteen terpenoids were detected at S2 (the stage of flower buds 1 day before full bloom), while twenty-two were detected at S3 (fully bloom flowers for 1 day). The data are shown in Table 1. The total ion chromatograms of scent components emitted from the flowers at two different developmental stages are shown in Figure 4. Linalool and \(\beta\)-pinene were the main compounds, accounting for 52.23% and 28.79% at S2. At S3, the contents of linalool and \(\beta\)-pinene accounted for 40.76% and 37.13% of the total compounds. In addition, there were three major components, D-limonene, \(\alpha\)-phellandrene, and \(\beta\)-phellandrene, which varied greatly between two stages. \({}^{1}\) Retention time (RT) in min. \({}^{2}\) Relative peak area expressed as percentage of the peak area of corresponding compound over the total peak areas of all identified volatiles. --, not detected. The relative contents of five major terpenoids were labeled in light gray. \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**No**} & **RT 1** & \multirow{2}{*}{**Terpenoids**} & \multicolumn{2}{c}{**Relative Content 2 (\%)**} \\ \cline{3-5}  & & & **S2** & **S3** \\ \hline
1 & 6.203 & \(\alpha\)-Thujene & 0.330 \(\pm\) 0.100 & 0.435 \(\pm\) 0.195 \\
2 & 6.374 & \(\alpha\)-Pinene & 0.665 \(\pm\) 0.065 & 0.990 \(\pm\) 0.210 \\
3 & 7.156 & \(\beta\)-Terpinene & 0.375 \(\pm\) 0.135 & 0.585 \(\pm\) 0.535 \\
4 & 7.577 & \(\beta\)-pinene & 28.865 \(\pm\) 1.095 & 37.000 \(\pm\) 2.830 \\
5 & 7.853 & \(\alpha\)-Phellandrene & 1.060 \(\pm\) 0.260 & 1.995 \(\pm\) 1.335 \\
6 & 8.011 & 2-Carene & — & 0.095 \(\pm\) 0.065 \\
7 & 8.162 & p-Cymene & 0.050 \(\pm\) 0.020 & 0.765 \(\pm\) 0.345 \\
8 & 8.287 & D-Limonene & 1.455 \(\pm\) 0.415 & 8.615 \(\pm\) 1.455 \\
9 & 8.333 & \(\beta\)-Phellandrene & 1.575 \(\pm\) 0.495 & 3.665 \(\pm\) 0.305 \\
10 & 8.491 & (_E_)-\(\beta\)-Ocimene & 0.130 \(\pm\) 0.020 & 0.300 \(\pm\) 0.140 \\
11 & 8.748 & \(\gamma\)-Terpinene & 0.210 \(\pm\) 0.040 & 0.150 \(\pm\) 0.070 \\
12 & 9.241 & Terpindene & 0.070 \(\pm\) 0.030 & 0.485 \(\pm\) 0.275 \\
13 & 9.609 & Linalool & 52.230 \(\pm\) 2.210 & 40.140 \(\pm\) 1.960 \\
14 & 10.010 & 2.4(10)-Thujadiene & — & 0.145 \(\pm\) 0.065 \\
15 & 10.885 & Terpinen-4-ol & 0.100 \(\pm\) 0.030 & 0.185 \(\pm\) 0.025 \\
16 & 11.102 & \(\alpha\)-Terpineol & 0.310 \(\pm\) 0.060 & 0.235 \(\pm\) 0.105 \\
17 & 14.481 & \(\alpha\)-Bergamotene & 0.025 \(\pm\) 0.015 & — \\
18 & 14.652 & trans-\(\beta\)-Farnesene & — & 0.085 \(\pm\) 0.075 \\
19 & 14.823 & cis-\(\beta\)-Farnesene & — & 0.185 \(\pm\) 0.085 \\
20 & 15.132 & (_Z_,_E_)-\(\pi\)Framesene & — & 0.170 \(\pm\) 0.080 \\
21 & 15.316 & \(\alpha\)-Farnesene & — & 0.205 \(\pm\) 0.065 \\
22 & 16.053 & cis-Nerolidol & — & 0.040 \(\pm\) 0.020 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Terpenoids identified from _Hosta_ flower volatiles of two different developmental floral stages using GC–MS analysis. All compounds were identified using the NIST database and quantified by comparing the peak area with the internal standard ethyl decylate. Data are presented as means (\(\pm\)SE, \(n\) = 3). The expression of Hs_TPS1_ and Hs_TPS2_ genes was consequently evaluated during petal development using quantitative RT-PCR to compare their transcription. The expression level of Hs_TPS2_ was much higher than that of Hs_TPS1_. The expression patterns of the two Hs_TPSs_ were higher at S2 and S3 compared with S1 and S4. Hs_TPS1_ expression was highest at S3, while Hs_TPS2_ expression was highest at S2.