# Effect of Mechanical Damage in Green-Making Process on Aroma of Rougui Tea
## 3 Results
### Analysis of DEGs and DEPs Related to Biosynthesis of Aroma Components
Some studies have shown that shaking and rocking can upregulate the LOX gene to increase the enzyme activity, thus promoting the biosynthesis of jasmine lactone, leading to the accumulation of volatile fatty acids [19]. In this study, the content of jasmine lactone in green-making also increased significantly (\(p<0.05\)). To summarize, the green-making and spreading process had significant effects on the \(\alpha\)-linolenic acid metabolic pathway, most of the related genes were upregulated, and the expression abundance of some proteins (AOS, OPCL, ACX) was higher than that in the transcriptional group, which was affected by the translation level's regulation or accumulation after transcription. This was beneficial to promote the synthesis of volatile aliphatic and jasmonic acid derivatives. Terpenoid aromatic compounds are the main substances that form the floral and fruity characteristics of RGT, among which monoterpene and sesquiterpene have the greatest influence. Most of the differential genes in this experiment were concentrated in the terpenoid backbone biosynthesis pathway (Figure 5B). This pathway can be divided into the mevalonate pathway (MVA) pathway and methylerythnitol 4-phosphate pathway (MEP) pathway, which are carried out in the cytoplasm and plastid, respectively [47]. In the cytoplasm, hydroxymethylglutaryl-CoA synthase (HMGS) is the rate-limiting enzyme of the MVA pathway [48]. Two genes regulating HMGS were extracted from the samples and upregulated in the process of green-making and spreading. The upregulation effect of green-making was obvious, and the trend of protein expression was the same. At the same time, 3-hydroxy-3-methyl glutaryl coenzyme A reductase (HMGR), the mevalonate kinase gene (MVK), the phosphomevalonate kinase gene (PMVK) and the methylglutarate diphosphate decarboxylase gene (MVD) in the MVA pathway were all upregulated during green-making, and they reached the highest levels at the end of green-making. The upregulation of related differential genes in LY was higher than that in CK. In the plastid, deoxyxylose-5-phosphate synthase (DXS) is a key rate-limiting enzyme in the regulation of monoterpene accumulation in the MEP pathway, and its genes are upregulated in Euphor-bia angustifolia. In addition, the 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR), 2-C-methyl-D-erythnitol-2,4-cyclodiphosphate synthase (ISPF), (E)-4-hydroxy-3-methylbut-2-enyl-diphosphate synthase (ISPG) and (E)-4-hydroxy-3-methylbut-2-enyl pyrophosphate reductase (ISPH) genes were upregulated under the mechanical injury and dehydration of green-making and spreading, and the upregulation was more significant in the CK group. ISPG and ISPH are key regulatory genes for the synthesis of monoterpene and sesquiterpenes' common precursors, dimethylallyl diphosphate (DMAPP) and isopentenyl diphosphate (IPP) [49]. It was found that the expression trend of the ISPG protein (TEA013763.1) was consistent with the transcriptome data. The expression of the 2-C-methyl-D-erythnitol-4-phosphate cytidyltransferase (ISPD) gene decreased gradually with green-making and spreading, while the expression of the 4-(cytidine-5'-diphosphate)-2-c-methyl-d-erythnitol kinase (ISPE) gene increased significantly at the first stage of green-making and spreading (\(p<0.05\)) and then decreased gradually. However, the expression of the isopentene diphosphate isomerase gene (IDI), which catalyzed the transformation of DMAPP and IPP, was upregulated, and the expression of two genes (TEA018159.1 and TEA030524.1) increased at the end of green-making. Six geranylgeranyl diphosphate synthase (GGPS) genes showed no obvious upregulation in the process of green-making; only one corresponding protein (TEA003988.1) was upregulated, and three genes encoding GGPS (TEA006109.1, TEA018068.1 and TEA019181.1) were significantly upregulated at the end of spreading (\(p<0.05\)). Geranyl-pp, synthesized by GGPS, is a precursor of monoterpene.