CLONING, EXPRESSION AND PURIFICATION CEL I ENDONUCLEASE ENZYME FROM CELERY PLANTS

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KHALIGHFARD S.; BANDEHPOUR M.; YASSAEE V.R.; PARIVAR K.; KAZEMI B.

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Paper Information

Journal: NEW CELLULAR & MOLECULAR BIOTECHNOLOGY JOURNAL Year:2012 | Volume:2 | Issue:8 Page(s): 79-87

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Title

CLONING, EXPRESSION AND PURIFICATION CEL I ENDONUCLEASE ENZYME FROM CELERY PLANTS

Author(s)

Keywords

CELI ENDONUCLEASE ENZYMESQ2

WESTERN BLOTTINGQ2

PLASMID PET32AQ2

RECOMBINANT PROTEINSQ2

AFFINITY CHROMATOGRAPHYQ2

Abstract

Aim and Background: CEL I, is isolated from celery and the first eukaryotic nuclease is known to have cleavage DNA with high specificity at sites of the two DNA strands in a heteroduplex DNA molecule. The aim of this study was cloning, expression and purification CEL I endonuclease and a simple method for the detection of mutations.Materials and Methods: CEL I mRNA was used for RT-PCR. CEL I cDNA was synthesized and cloned into T-vector pTZ57R, then CEL I subcloned into pET32a expression vector. Recombinant plasmid was transformed into BL2I bacterial cells. Expression of recombinant plasmid was analyzed by western blot technique. Protein was purified by AFFINITY CHROMATOGRAPHY. In first study, Recombinant enzyme was active in mutation detection in Product PCR normal and mutant BRCA1 gene.Results: The recombinant (pET32a/CEL I) was successfully expressed in BL2I cells. Western blot analysis showed that the successfully expressed CEL I in the cells transfected. The first study showed that recombinant enzyme is known to nick mismatch sites of the two DNA strands in a heteroduplex DNA molecule.Conclusion: The expression of CEL I in BL2I cells was strong. In vitro, purification protein was successful. Recombinant nucleases CEL I known to nick mismatch sites in a heteroduplex DNA molecule.

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APA: Copy

KHALIGHFARD, S., BANDEHPOUR, M., YASSAEE, V.R., PARIVAR, K., & KAZEMI, B.. (2012). CLONING, EXPRESSION AND PURIFICATION CEL I ENDONUCLEASE ENZYME FROM CELERY PLANTS. NEW CELLULAR & MOLECULAR BIOTECHNOLOGY JOURNAL, 2(8), 79-87. SID. https://sid.ir/paper/204504/en

Vancouver: Copy

KHALIGHFARD S., BANDEHPOUR M., YASSAEE V.R., PARIVAR K., KAZEMI B.. CLONING, EXPRESSION AND PURIFICATION CEL I ENDONUCLEASE ENZYME FROM CELERY PLANTS. NEW CELLULAR & MOLECULAR BIOTECHNOLOGY JOURNAL[Internet]. 2012;2(8):79-87. Available from: https://sid.ir/paper/204504/en

IEEE: Copy

S. KHALIGHFARD, M. BANDEHPOUR, V.R. YASSAEE, K. PARIVAR, and B. KAZEMI, “CLONING, EXPRESSION AND PURIFICATION CEL I ENDONUCLEASE ENZYME FROM CELERY PLANTS,” NEW CELLULAR & MOLECULAR BIOTECHNOLOGY JOURNAL, vol. 2, no. 8, pp. 79–87, 2012, [Online]. Available: https://sid.ir/paper/204504/en

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