Comparative Evaluation of Expression Vectors (pET32a and pET25b) in Expression of Polyepitopic Sequence Synthesized from Leishmania infantum Antigens

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TAHERZADEH M.; FOULADVAND M.A.; KAZEMI B.

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Journal: IRANIAN SOUTH MEDICAL JOURNAL (ISMJ) Year:2020 | Volume:23 | Issue:2 Page(s): 143-152

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Title

Comparative Evaluation of Expression Vectors (pET32a and pET25b) in Expression of Polyepitopic Sequence Synthesized from Leishmania infantum Antigens

Author(s)

TAHERZADEH M. | FOULADVAND M.A. | KAZEMI B. | Issue Writer Certificate

Keywords

Poly-epitopic structureQ1

Leishmania infantumQ2

Expression levelQ1

pET25b vectorQ1

pET32a vectorQ1

Abstract

Background: Molecular methods are nowadays used to diagnose diseases, produce vaccines, drugs and recombinant proteins. Therefore, one of the important steps in these procedures is selecting the appropriate vector for cloning and expression of the target genes. Because the expression of synthetic polygenes or so-called chimeric antigens with high molecular weight requires special conditions, the aim of this study was to evaluate the importance of vector type for expression of such antigens. Materials and Methods: The 1250bp sequence consisting of epitopes from eight important antigens of Leishmania infantum parasite was designed and synthesized by Biomatic Company (Cambridge, Canada). The sequence was cloned separately in two expression vectors pET25b and pET32a and then transformed into E. coli BL21 (DE3) and expressed under similar appropriate conditions. Bacterial lysates were analyzed by SDS PAGE and evaluated by Western blotting. Results: of SDS-PAGE and Western blot analysis showed that intact recombinant protein production by pET25b vector was not successful. However, the recombinant protein resulted from the expression of the aforementioned poly-epitope in pET32a vector was successfully produced and confirmed. Conclusion: According to the successful expression confirmation of the poly-epitopic sequence within the pET32a vector and furthermore the failure to obtain protein in the pET25b vector showed that in the case of some specific mosaic sequences expression, protein isolation will be difficult because of low solubility. So, the expression vector choose should be made more carefully.

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APA: Copy

TAHERZADEH, M., FOULADVAND, M.A., & KAZEMI, B.. (2020). Comparative Evaluation of Expression Vectors (pET32a and pET25b) in Expression of Polyepitopic Sequence Synthesized from Leishmania infantum Antigens. IRANIAN SOUTH MEDICAL JOURNAL (ISMJ), 23(2 ), 143-152. SID. https://sid.ir/paper/361544/en

Vancouver: Copy

TAHERZADEH M., FOULADVAND M.A., KAZEMI B.. Comparative Evaluation of Expression Vectors (pET32a and pET25b) in Expression of Polyepitopic Sequence Synthesized from Leishmania infantum Antigens. IRANIAN SOUTH MEDICAL JOURNAL (ISMJ)[Internet]. 2020;23(2 ):143-152. Available from: https://sid.ir/paper/361544/en

IEEE: Copy

M. TAHERZADEH, M.A. FOULADVAND, and B. KAZEMI, “Comparative Evaluation of Expression Vectors (pET32a and pET25b) in Expression of Polyepitopic Sequence Synthesized from Leishmania infantum Antigens,” IRANIAN SOUTH MEDICAL JOURNAL (ISMJ), vol. 23, no. 2 , pp. 143–152, 2020, [Online]. Available: https://sid.ir/paper/361544/en

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