Cloning and Expression of 2S Albumin As a Major Allergen of Persian Walnut

Autor:	Maryam Vakili Moghadam, Fereshteh Sadat Rasi Varaee, Reza Jafari, Kobra Mokhtarian, Reza Falak, Mehran Gholamin, Mohsen Mohammadi
Rok vydání:	2020
Předmět:	Cloning Chemistry medicine.disease_cause Molecular biology law.invention Blot Allergen Metal affinity chromatography law Recombinant DNA medicine 2s albumin Vector (molecular biology) Target protein
Zdroj:	Immunoregulation. :51-60
ISSN:	2588-6088 2588-5200
DOI:	10.32598/immunoregulation.3.1.6
Popis:	Background: Food hypersensitivity to walnut usually results in mild symptoms; however, several cases of anaphylactic reactions to this product have been observed. This study aimed to determine the immunochemical characteristics of the Persian walnut and provide the recombinant form of its main allergen. Materials and Methods: The allergenic proteins of the Persian walnut were extracted by standard procedure. The IgE-binding profile was determined by common immunoassays, including ELISA and Western blotting. The characteristics of the main allergenic protein which showed a stronger and higher frequency of IgE-reactivity with the patient’s sera was identified by MALDI-TOF-TOF method. The conventional PCR was used for the amplification of the coding sequence of the target protein which was then inserted into pET-21b(+) vector and expressed in E. coli BL21. The recombinant allergen was purified by metal affinity chromatography and the ELISA and immunoblotting assays were used for the evaluation of the IgE-binding capacity of the recombinant protein.Results: All patients showed a considerable specific IgE-reactivity to total extract (OD 0.58±0.43 versus 0.047±0.026; P
Databáze:	OpenAIRE
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