RESUMO
Background: The homologous recombination [HR] is one of the conventional cloning methods for the production of recombinant DNA. It is a quick method for in vivo DNA cloning without using the high cost restriction enzymes. A few modifications in the cloning procedure can increase the efficiency of this method.
Materials and Methods: In this study, effect of heating on the rate of the IgG1 heavy chain gene cloning was investigated in the HR method and then it was compared with HR method without heating and traditional cloning method. For doing this comparison, three cloning methods including HR, HR with the heat treatment, and traditional cloning were used to clone the human IgG1 heavy chain into the pcDNA3.1[+] plasmid.
Results: The results showed that adding heat in the HR method converts insert and vector from the double strand DNA to the single strand DNA. This allows them to copulate with each other better and faster than the two other methods. The heat addition also helps the cell enzyme system for a faster and easier recombination and moreover it increases the cloning efficiency of the HR method in case of in vitro processing.
Conclusion: The results showed that giving heat in the HR method increases cloning rate 7.5% and this increase reaches 10% in comparison with the traditional method.
RESUMO
Prevalence of hepatitis B infection, existence of non-responder populations and the weakness of the classic Alum adjuvant in inducing proper cellular immune responses display the importance of designing more efficient vaccines. Many studies have demonstrated interactions between iron and the immune system. In the present study, we have evaluated the effects of two iron containing structures, ferrous sulphate and AJc-F nano complex on immune response. In this experimental study, female Balb/c mice were injected 3 times with hepatitis B vaccine. The proliferative response of the splenocytes was evaluated using bromodeoxyuridine assay and interleukin-4, interferon-gamma [IFN-gamma] levels and total specific antibodies were examined by the ELISA method. Data analysis was done by SPSS software version 18. AJc-F nano complex significantly increased total antibody and cytokine levels compared to ferrous sulfate and control groups. Ferrous sulfate severely decreased total antibody and IL-4 levels compared to control group. Both compounds increased splenocyte proliferation and IFN-gamma levels compared to control group. The results show that AJc-F, as an iron-containing nano complex, could efficiently increase humoral and cellular immune responses against hepatitis B vaccine