## 2 Results and Discussion
### Relationship between C6 and C9 Aldehydes and Gene Expressions in the 9930 Line  
In order to explore potential correlations between aldehydes, the folds of _CsLOXs_ and _CsHPLs_ gene expression patterns were used for HCA analysis, and N-deficient treatments were compared to N-sufficient treatments. HCA data resulted in two distinct sample clusters (Figure 6). Cluster I included six _CsLOXs_ (_CsLOX2_, 9, 17, 19, 20, and 22) that displayed higher up-regulated expression patterns during N-deficient treatments. Cluster II included all nine aldehydes, six _CsLOXs_, and two _CsHPLs_. The expression of _CsHPLI_ was significantly increased at 5 d, meanwhile the amount of (_E_)-2-hexenal and hexanal were also significantly up-regulated. This result indicated that _CsLOX2_, 9, 17, 19, 20, and 22 were responsive to N-deficiency and had higher up-regulated expression patterns during short-term N-deficient treatments, thus the increase in C6 aldehydes might be induced by the up-regulation of _CsHPLI_. Changes of C6 and C9 Volatile Content during N-Deficient Treatment in Six Other Inbred Cucumber Lines  
In order to further study the response of the C6 and C9 volatile metabolism to N-deficient treatments, six other inbred cucumber lines were cultivated in N-deficient nutrition solution and sampled at Day 5 for detecting leaf volatiles. Seven short-chain leaf volatiles (hexanal, (_E_,_E_)-2,_A_-hexadienal, (_E_)-2-hexenal, nonanal, (_Z_)-6-nonenal, (_E_,_Z_)-2,6-nonadienal, and (_E_,_Z_)-2,6-nonadienol) were detected in cucumber leaves, as shown in Figure 7 and Table S3, compared to N-sufficient treatments, the total amount of short-chain volatiles of all six lines was significantly improved in the N-deficient treatments, up-regulation of 0.87- to 1.96-fold was displayed, and the amount of C6 volatiles of all the six lines and C9 volatiles of five lines increased. Of the six inbred lines, (_E_)-2-hexenal was still the main component, displaying as much as 75.38% in all treatments. (_E_,_Z_)-2,6-nonadienol was the smallest component, displaying between only 0.02% and 0.73% in all treatments. During N-deficient treatments, changes in volatile components were observed among six inbred lines: 32 volatiles were up-regulated and 10 volatiles were down-regulated at 5 day. All of the (_E_)-2-hexenal amounts, which had green-like flavor, as well as (_E_,_Z_)-2,6-nonadienal, which had cucumber-like flavor, were improved compared to N-sufficient treatments. The other five volatiles displayed down-regulation in some lines. Among the six inbred lines, different responses to N-deficiency were also observed. For A37 and A91, the amount of all seven volatiles increased. For A74, only one volatile decreased, but for A23, the content of (_E_)-2-hexenal, (_E_,_Z_)-2,6-nonadienol, and (_E_,_Z_)-2,6-nonadienal displayed up-regulation, whereas the amount of the other four volatiles decreased. Figure 5: Changes in content of C9 aldehydes in 9930 leaves during N-sufficient and deficient treatments. Bars present means \(\pm\) standard deviations. \({}^{\star}\) and \({}^{\star\star}\) mean significant difference from control at \(p<0.05\) and \(p<0.01\), respectively. N+ and N – mean nitrogen sufficient and nitrogen deficient treatments, respectively. Values are means of three replicates. Combined with the changes of 9930 and the volatile content of six other inbred lines, we suggest that the amount of short-chain leaf volatiles in cucumber seedlings could be significantly improved in response to nitrogen deficiency. A previous study showed that reactive short-chain leaf volatiles  
Figure 6: Hierarchical cluster dendrogram and heat map of volatiles content and gene expression folds during N-sufficient and deficient treatments in 9930 leaves. Leaves were sampled at 1, 2, 3, 4, 5, 6 and 7 day after N-deficient treatments. The heat map has been generated based on the fold-change values of volatile contents, _CsLOXs_ and _CsHPL_ expression level in the treated samples when compared with its unstressed control samples by MetaboAnalyst 3.0 software.