## 3 Results
### Genomic and Expression Analyses
#### 3.1.3 Polymerase Chain Reaction (PCR)  
After PCR of cDNA with primers MG82 (5'-CTGGGAGCTG-3'), two amplicons (not present in the JA genotype) could be observed in the JB genotype (Figure 4).  
The fragments isolated were re-amplified. Only seven reproducible fragments were considered for sequencing and were searched against the EMBL database using the BLAST program. Two sequenced cDNAs showed similarity with known activity sequences of enzymes aldehyde dehydrogenase and class III peroxidase, whereas the other sequences showed homology with expressed products involved in the flavonoid pathway. The application of RAPD markers produced a high number of bands and showed the greatest potential to discriminate polymorphic DNA segments. The homologies founded (aldehyde dehydrogenase, class III peroxidase, and genes involved in the flavonoid pathway) represented the first step in investigating the metabolic pathways involved in the biosynthesis of secondary metabolites. Genes in the aldehyde dehydrogenase (ALDH) family play a role in the metabolism of many molecules, such as cholesterol, fatty acids, amino acids, and ethanol. Furthermore, ALDH enzymes detoxify substances such as alcohol, pollutants, and toxins produced within cells. One of the most important pathways in aldehyde metabolism is their oxidation to carboxylic acids by ALDHs. The oxidation of the carbonyl group was considered as a positive detoxification process;  
Figure 4: Amplification of cDNA of genotype A (JatA) and B (JatB) obtained from _J. curas_ applying MG82 primer. Ladder = DNA molecular size marker; CTRL– = negative control (no cDNA); CTRL+ = positive control (cDNA that has been successfully used in previous PCRs).  
Figure 3: Amplification of genomic DNA of _J. curas_ obtained applying MG108 primer.  
however, in humans the polymorphism (i.e., the presence of more alleles of the same gene) of the ALDH gene is correlated to the occurrence of phenotypic diseases or metabolic disorders [24]. Class III peroxidase is involved in key processes that determine the architecture and the defense properties of the plant cell wall and its role in stress resistance and adaptation, as well as in auxin catabolism, in secondary metabolism and in hydrogen peroxide scavenging. In vitro studies have shown the capability of plant peroxididases to accept as substrates a number of vacuolar metabolites (i.e., phenols, flavonoids, and alkaloids), and it has been suggested that peroxididases play a specific role in the metabolism of these compounds in vivo [25; 26]. The flavonoids show a variety of pharmacological activities such as antimicrobial, hepatoprotective, anti-inflammatory, anti-oxidant, anticancer, and antiviral properties [27]. Because flavonoids have been found in _J. curcas_, the evaluation of the expression of genes involved in their pathway, using specific primers [17], could represent an important aspect in view of a possible medical use of this plant. Finally, the results obtained for the homology analysis highlighted some sequences for which the E-value of the BLAST homologue analysis was more than 1e-50, suggesting that they are novel genes.