利用DNA分子指纹图谱技术检测小麦粉中害虫的研究.docVIP

河南工业大学硕士学位论文 PAGE PAGE 1 PAGE PAGE 12 国家公益行业专项201513002-05 收稿日期-2018-01-26 作者简介：鲁玉杰 女 1971年9月出生 教授 粮食储藏与品质控制 利用DNA分子指纹图谱技术检测小麦粉中害虫的研究 鲁玉杰 袁园 刘凤杰 王争艳 何向楠 （河南工业大学粮油食品学院， 河南省小麦储藏协同创新中心 ，郑州，450001） 摘要 为开发快速检测我国成品粮中害虫的技术，研究了DNA分子指纹图谱技术检测小麦粉中主要害虫的碎片和虫卵的含量。结果表明，通过对害虫的延长因子EF1基因片段引物的筛选和PCR条件的选择，确定了有效地检测出来小麦粉中的赤拟谷盗、杂拟谷盗和锯谷盗的成虫碎片、幼虫和卵通用引物和PCR扩增条件。通过改良的PCR技术可在4 h内快速检测杂拟谷盗、锯谷盗的卵、幼虫和成虫碎片。最低检出限为可检测出小麦粉中0.05%（w/w）卵和幼虫、1%成虫的碎片，灵敏度超过了国际标准。DNA指纹图谱中电泳条带亮度（Y）与被检测的不同害虫碎片含量（X）成正相关，回归方程为Y=858.36X+74.540相关系数r为0.961 9。利用该技术检测随机取样的小麦粉中成虫的碎片，检测的准确率最高可达80.56%。结果说明了利用DNA分子指纹图谱技术可快速检测出小麦粉中害虫的碎片、幼虫和卵含量，并可预测小麦粉中害虫的感染程度。本研究为开发快速小麦粉中害虫的快速检测技术具有重要的参考意义。 关键词 DNA分子指纹图谱 害虫碎片 赤拟谷盗 杂拟谷盗 小麦粉 最低检测限 中图分类号：TS201.6;Q969.98 文献标识码：A 文章编号：1003-0174（ ） - - DNA Molecular Fingerprint Pattern Technology to Detect Pests in Wheat Flour Lu Yujie, Yuan Yuan, Fengjie Liu, Zhengyan Wang, Xiangnan He (School of Food Science and Technology，Henan University of Technology, Zhengzhou, Henan Province, 450001) Abstract To develop the technology of fast detection of insect in our grain, the molecular fingerprint technique was used to detect quickly adults fragments and eggs of insect in wheat flour. The results showed that the universal primers and condition of PCR were confirme that could be detected the adult fragments, larval and eggs of Tribolium castaneum ,T. confusum and Oryzaephilus surinamensis by selecting the primers for insect elongation factor gene EF1 and PCR condition. Improved PCR technology could quickly detect the eggs、larva and fragments of T. confusum and O. surinamensis in wheat flour in 4hours.The lowest detection limitation for eggs and larvae of wheat flour is 0.05% (w/w), for adult fragment is 1%(w/w). Sensitivity of detection exceeds the international standard. There was a positive correlation brightness of DNA agarose gel electrophoresis banding (Y) and adult fragment content in flour(X) and the regression equation is Y=858.36X+74.540 (r= 0.9619). The highest prediction accu