脱氧雪腐镰刀菌烯醇的环七肽模拟表位筛选  

Screening of the mimic epitopes of deoxynivalenol from cyclo-heptapeptide library

作　　者：余鹃 贾彦琼 骆敏儿 王宏[1] 

机构地区：[1]暨南大学生命科学技术学院生物工程学系,广东广州510632

出　　处：《食品工业科技》2017年第24期133-136,147,共5页Science and Technology of Food Industry

基　　金："十一五"国家科技支撑计划项目(2008BAK42B05)

摘　　要：利用噬菌体随机环七肽库筛选脱氧雪腐镰刀菌烯醇(Deoxynivalenol,DON)模拟表位。以DON单抗为靶分子,四轮固相淘选噬菌体随机环七肽库,筛选出阳性噬菌体,并通过ELISA实验检测其特异性。对阳性克隆的DNA进行测序,取序列不同的阳性克隆,分别建立阳性噬菌体与DON标准品的竞争抑制曲线。对30个噬菌体单克隆进行活性测定,结果显示能与DON单抗结合的噬菌体克隆有28株,其中的22株能被DON标准品阻断与DON单抗结合。DNA测序结果显示,21株阳性噬菌体的序列是PFPNHPY,另一株的序列是TPWTQHL。其相应噬菌体与DON毒素标准品的竞争曲线结果显示,8号噬菌体建立的竞争抑制曲线线性范围是0.0292~0.636μg/mL,IC_(50)为0.169μg/mL;4号噬菌体建立的竞争抑制曲线线性范围是0.0124~0.271μg/mL,IC_(50)为0.058μg/mL。在随机环七肽库中筛选到TPWTQHL和PFPNHPY两个DON模拟表位,可替代DON毒素标准品,建立DON的免疫学无毒检测技术。Screening of mimetic epitopes of deoxynivalenol by using random Ph.D-7 library.DON monoclonal antibody as the target molecule, and Ph.D-7 library had been panned for 4 times. The positive phage was screened out and tested by ELISAspeeificity.The activity of 30 phage monoclones was determined by indirect ELISA and indirect competitive ELISA. It showed that 28 phages could combined with DON monoelonal antibody and 22 of them were able to compete with DON standards for binding to DON mAbs.The results of DNA sequencing showed sequence of 2l was PFPNHPY, and the another one was TPWTQHL.The competition curve of corresponding phage and DON toxin standard product showed inhibition curve for the linear range of NO.8 phage was 0.0292-0.636 I^g/mL and IC^0 was 0.169 p^g/mL,and NO.4 phage was 0.0124-0.271 p,g/mL, ICs0 was 0.058 p^g/mL.TPWTQHL and PFPNHPY were obtained from Ph.D-7 phage display peptide library.The competitive inhibition curves of phage and DON were established respectively.Preliminary validation shows that the DON toxin standard can be replaced by two DON mimotopes and can be established DON immunological non-toxic detection techniques.

关 键 词：脱氧雪腐镰刀菌烯醇 噬菌体展示技术 模拟表位 

分 类 号：TS201.3[轻工技术与工程—食品科学]