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Construction of Naive Channel Catfish ScFv Phage Display Antibody Library and Preliminary Identificiation

Author: WangHui
Tutor: ZouShiPing
School: Huazhong Agricultural University
Course: Aquaculture
Keywords: Channel Catfish Phage antibody library Specific antibodies Filter
CLC: R392
Type: Master's thesis
Year: 2011
Downloads: 6
Quote: 0
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Abstract


The antibody has an important role in the study of biology , phage display technology found antibodies . Antibody library technology research and application of aquatic organisms on basic or blank , build Ictalurus natural single-stranded phage antibody library research . In this study, using non - immunized channel catfish , from its lymphocyte cell extract RNA, RT-PCR amplified VH and light chain variable regions of the antibody heavy chain variable region the VL gene purified recycled VH and VL , first use Linker fragment - containing primers will Linker fragment connected to the VL gene segment up into a single-chain antibody gene ( scFv ) , and then assembled and VH fragment scFv gene fragment after SfiI sites with restriction endonuclease Notl primer amplification , Get the objective fragment of about 1300bp . Then the fragment purified SfiI , NotI digest and transform Escherichia coli competent cells after SfiI, NotI digested vector pCANTAB-5E connection . The transformed bacteria obtained after secondary the phage VCSM13 role Ictalurus natural single-stranded phage antibody library . Extraction of plasmid by PCR identification to prove that the exogenous fragment was inserted into , and measured capacity of 5.8 × 105, indicating the success of building a database . Successfully build the Channel Catfish natural single chain phage antibody library 's foundation , OVA-SMZ antigens are building spots fork tail channel catfish natural single chain phage antibody library carried the adsorption of three \set the selection process , and access to the OVA-SMZ secondary phage antibody library . Select a single colony from the antibody library phage antibody prepared for ELISA identified to achieve the expected results , proved once again that building a database successfully . Completion of this study , on the one hand, the antibody preparation and study of new biological products to explore new avenues , there are important ; drug residue detection research opens a new direction for the diagnosis and treatment of diseases of aquatic organisms .

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CLC: > Medicine, health > Basic Medical > Medical Immunology
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