转基因玉米MON87427梯度含量基体标准物质的研制  

Development of A Set of Matrix Reference Materials in Different Mass Fractions of Genetically Modified Maize MON87427

作　　者：李俊[1] 单露英 肖芳 李允静[1] 高鸿飞 翟杉杉 吴刚[1] 张秀杰 武玉花[1] LI Jun;SHAN LuYing;XIAO Fang;LI YunJing;GAO HongFei;ZHAI ShanShan;WU Gang;ZHANG XiuJie;WU YuHua(Oil Crops Research Institute,Chinese Academy of Agricultural Sciences/Key Laboratory of agricultural genetically modified organism traceability,Ministry of Agriculture and Rural Affairs,Wuhan 430062;Development Center of Science and Technology,Ministry of Agriculture and Rural Affairs,Beijing 100025)

机构地区：[1]中国农业科学院油料作物研究所/农业农村部农业转基因生物溯源重点实验室,武汉430062 [2]农业农村部科技发展中心,北京100025

出　　处：《中国农业科学》2023年第8期1444-1455,共12页Scientia Agricultura Sinica

基　　金：转基因生物新品种培育重大专项(2016ZX08012-003)。

摘　　要：【目的】转基因检测标准物质是转基因生物安全监管、标识制度实施的物质基础,是实验室内检测数据溯源、实验室间检测数据可比的保证。中国于2017年批准进口转基因玉米MON87427用作加工原料,其安全监管和检测急需研制有证标准物质。【方法】以开发商提供的转基因玉米MON87427杂合种子和非转基因受体种子为原材料,对转基因玉米MON87427和其受体种子分别清洗、干燥、冷冻研磨、粒径及含水量检测,按质量分数(以干基计)称量配制得到质量分数分别为60.0、99.5和1000.0 mg·g^(-1)的转基因玉米MON87427基体标准物质MON87427a、MON87427b和MON87427c。运用实时荧光定量PCR进行粉末的均匀性初检,初检合格后分装。用MON87427/zSSIIb二重微滴数字PCR(ddPCR)方法进行标准物质均匀性、稳定性评估和8家实验室联合定值。根据《标准物质定值的通用原则及统计学原理》(JJF 1343)进行标准物质均匀性评估、稳定性评估、联合定值数据的处理及不确定度评定。【结果】本批标准物质有MON87427a、MON87427b、MON87427c 3个浓度,粒径小于200μm的粉末大于80%,含水量低于5%。用棕色玻璃瓶分装,不低于1.0 g/瓶,充氮后轧盖,3个浓度的标准物质各分装400瓶。3种标准物质均匀性检验的F值均小于临界值F0.05(14,30)(2.04),在瓶内和瓶间具有良好的均匀性,使用时,最小取样量为100 mg。标准物质在25℃、37℃和60℃条件下可稳定储存14 d,表明在常温下运输14 d标准物质特性量值不发生趋势性变化;在4℃和-20℃条件下长期稳定性达到12个月;开盖取样5次,标准物质特性量值未发生显著偏离。8家实验室联合测定的标准物质拷贝数比值无异常值、呈正态分布、且等精度,标准物质MON87427a、MON87427b、MON87427c的标准值及扩展不确定度分别为(2.92±0.44)%、(4.89±0.57)%、(52.1±3.4)%。【结论】研制的转基因玉米MON87427梯度含量基体�【Objective】The GMO(genetically modified organism)reference materials(RMs)are the material basis for GMO safety supervision and labeling policy implementation.During GMO detection the utilization of RMs guarantees the traceability of quantitative results within laboratory and the comparability of quantitative results between laboratories.GM(genetically modified)maize MON87427 has been approved to be imported as raw material in China,it’s urgent to develop certified reference materials(CRMs)for safety supervision and quantification.【Method】The GM maize MON87427 hybrid seeds and non-GM counterparts provided by the developer were used as raw materials to perform washing,drying,freeze-grinding,particle size measurement,and moisture content measurement,sequentially.The matrix RMs of MON87427a,MON87427b and MON87427c were produced by blending the seed powder of the GM maize MON87427 and a non-GM counterpart in GMO mass fractions of 60.0 mg·g^(-1),99.5 mg·g^(-1) and 1000.0 mg·g^(-1) on a dry basis.The real-time quantitative PCR was used to conduct an initial assessment of homogeneity before packing.The MON87427/zSSIIb duplex digital PCR(ddPCR)was used to evaluate the homogeneity and stability of the RMs as well as the collaborative characterization by 8 qualified laboratories.The data processing of homogeneity,stability,collaborative characterization together with uncertainty evaluation of RMs were carried out according to the standard"General and Statistical Principles for Characterization of Reference Materials"(JJF 1343).【Result】This batch of RMs contains three RMs of MON87427a,MON87427b,and MON87427c in different mass fractions,with more than 80%of particle size of less than 200μm and less than 5%of moisture content.The RMs were packed in brown glass bottles,nitrogen flushed before capping,bottling amount was not less than 1.0 g/bottle,a total of 400 bottles were produced for each mass fraction RM.The calculated F values of the homogeneity test were all less than the critical value of F0.05(14,30)(2

关 键 词：转基因玉米MON87427 基体标准物质 二重微滴数字PCR 特性量值 不确定度 

分 类 号：S513[农业科学—作物学]