RESUMEN
Objective: To screen for the optimal qPCR primers for Echinococcus multilocularis apomucin gene (Em-apo) and analyze Em-apo expression. Methods: Primers were designed based on 4 Em-apo sequences from GeneDB. Primer specificity and PCR efficiency were determined, based on which the optimal primer pairs were selected. Alterations of Em-apo expression in 1 000 E. multilocularis protoscoleces treated with albendazoleï¼5 µg/mlï¼ and insulinï¼100 ng/mlï¼ were separately assessed using the selected primers. DMSO used in albendazole dilution and in PBS insulin dilution were used as the control. Results: Specific primers for Em-apo-1, Em-apo-2/3, Em-apo-4 and actin were selected. qPCR melting curves revealed a single peak for each primer pair and an amplification efficiency from 95% to 101%. The qPCR showed increased expression of Em-apo-1ï¼1.51±0.27ï¼, Em-apo-2/3 ï¼1.39±0.30ï¼ and Em-apo-4ï¼1.14±0.18ï¼ after albendazole treatment in comparison to the DMSO controlï¼1.00ï¼ï¼P>0.05 among the three genesï¼; and an unaltered Em-apo-1 expression, slightly decreased Em-apo-4 expression, and significantly decreased Em-apo-2/3 expressionï¼0.73±0.09ï¼ after insulin treatment in comparison to the PBS control (P>0.05 among the three genesï¼. Conclusion: The selected specific primers for Em-apo genes can be used to analyze the gene expression by qPCR. Treatment with albendazole and insulin show certain effects on the expression of Em-apo genes in E. multilocularis protoscoleces.
