Discussion
The aim of our study was the characterization of the expression of  A. albimanus zpg  (AALB006050) and boule (AALB005972orthologues genes, in order to determine if they could be considered as targets for genetic approaches to induce sterility in this species. For this, processes of RNA extraction, cDNA synthesis and qPCR were standardized. 
At the beginning, RNA extraction presented some difficulties due to the big difference that exists between life stages in insects with developmental hemimetabolism, like An. albimanus. The protein synthesis and macromolecules composition at each stage may be a source of variation of this process, and reason for this make the number of individuals in each pool of extraction vary. The RNA obtained had an absorbance values at 260/280 nm between 2.0-2.2, which is generally accepted as pure RNA samples. After the RNA extraction, all the samples were treated with Dnase. Genomic DNA  could lead to low Ct values in the qPCR and non-specific amplifications.  As a quality control, it was necessary  to make conventional PCR after DNase treatment and cDNA synthesis, to prevent traces of genomic DNA or to be sure that the sample only contains cDNA. Followed cDNA synthesis, the samples were diluted 10 times fold, aliquoted and frozen at -70 C. Primer efficiency for qPCR analysis was calculated by using concentrations of 0.4 and 0.6uM for each pair of primers at pupae and adult stages. Finally, primers of 0.4uM were used because they efficiency values were closer to the optimal values (90-110%) than 0.6uM primers (Table No. 1). 
Relative expression graphs of boule and zpg are shown in Fig. 1 to 6. Actin gene was used as the housekeeping gene in all analyses.  Figure 1 and 4 presented that orthologues of boule and zpg are expressed similarly through out all larval stages. In Figure 2 and 5, relative expression in pupa is observed, and there is no significant difference between male and female among the genes analyzed. In male and female adults,  boule showed the same levels of relative expression. This fact could be attributed that BOLL protein may have the same levels of expression in gonads and somatic tissues in both sexes, as it was reported by SekinĂ© et al (2015)Interestingly, zpg orthologue (AALB006050) is expressed  70- times fold higher in females than males of adults An. albimanus. This result is presented in figure 6.  Inexin 4, or zpg, is well known in Drosophila as a  gap-juction protein. ZPG has been reported necessary for germ cells differentiation in gametogenesis of both sexes in fly (Tazuke et al 2002). In other organisms such as Bactrocera dorsalis, it was reported that this gene has higher expression in testis than ovaries (War Ali et al 2017). 
Calkins et al (2015) and Whyard et al (2015) reported that zpg was no detectable in larvae L4 or pupae. Those results agrees with our findings in larvae and pupae An. albimanus. The over-expression of zpg orthologue (AALB006050) gene in adult females, could lead to the assumption that  a higher demand of the ZPG protein sythtesis is more necessary in females than in males, but the cellular pathways and function still remained unclear. This finding would make this gene a prime candidate for further studies aiming to use genetic approaches to eliminate the vector. In order to know how much these genes are specifically expressed, it is necessary to dissect tissues of An. albimanus. Noticeable, the genes analyzed may had changed in the evolution pathways which may be the reason that these results differ with the results reported in other species. 
To conclude, it is necessary to carry out another studies in which characteristic relative expression of these genes are determined. Likewise, it is feasible to expect that the silencing of AALB006050 (zpg) gene by RNA interference will have consequences in females. We need to analyzed where boll is specifically expressed to infer wether it can be used as a candidate gene to be silenced in male An. albimanus as it was made in Ae. aegypti. The analysis of variability of these genes whithin the Anopheline, both in sequence and function, may generate important information to create efficient and specific tools for the control of vector-borned diseases like malaria.