*2.7. Gene Expression Analysis qRT-PCR Validation*

The expression levels of the *CsLEA 14*, *CsLEA 19*, *CsLEA 37* and *CsLEA 38* genes were induced after 24 h of ABA or heat treatment but were not tissue specific (Figure 5). To validate results from expression profile analysis, qRT-PCR was carried out for *CsLEA 14*, *CsLEA 19* (from the *LEA\_2* subfamily) and for *CsLEA37* and *CsLEA 38* (from the *Dehydrin* subfamily) as these genes showed a relatively high number of stress related *cis* acting elements and motifs.

We carried out qRT-PCR analyses using *CsLEA14* and *CsLEA19* (*LEA\_2* subfamily), and *CsLEA37* and *CsLEA 38* (*Dehydrin* family). Results showed that after 24 h heat treatment the expression levels of these four genes up-regulated by 156.5 (*CsLEA 19*), 95.8 (*CsLEA14*), 52.6 (*CsLEA38*) and 14.6 fold (*CsLEA37*) in *C. songorica* leaves compared with the untreated plant leaf samples. The expression levels of these four genes were slightly up-regulated after the ABA treatment, especially *CsLEA 38* (14.6 fold, Figure 6).

**Figure 4.** Locations of the 44 *CsLEA* genes on 15 chromosomes of *C. songorica*.

**Figure 5.** Hierarchical clustering of *CsLEA* gene expression profiles in root and shoot tissues after 24 h heat or ABA treatment. The log transformed values for the relative expressions of *CsLEA* genes were used for the hierarchical clustering analysis. The blue scale means low transcript expression and the red scale means high transcript expression.

**Figure 6.** Hierarchical clustering of *CsLEA* gene expression profiles in root and shoot tissues after 24 h heat or ABA treatment. The log transformed values for the relative expressions of *CsLEA* genes were used for the hierarchical clustering analysis. The blue scale means low transcript expression and the red scale means high transcript expression.
