限制性内切酶酶切确证河豚鱼成分PCR检测结果  

Confirmation of PCR Results for Puffer Fish Components by Restriction Endonuclease Digestion

作　　者：曲良苗[1,2] 陈文炳[1,3] 缪婷玉[1,3] 邵碧英[1,3] 彭娟[1,3] 江树勋[1,3] 

机构地区：[1]福建出入境检验检疫局检验检疫技术中心,福建福州350001 [2]福建农林大学食品科学学院,福建福州350003 [3]福建省检验检疫技术研究重点实验室,福建福州350001

出　　处：《食品科学》2014年第8期169-173,共5页Food Science

基　　金：国家质量监督境检验检疫总局科技计划项目(2008IK175;2013IK149)

摘　　要：动植物成分的聚合酶链式反应(polymerase chain reaction,PCR)检测往往出现假阳性结果,为了有效排除河豚鱼成分检测中出现的假阳性现象,提高检测结果的准确性,应用河豚鱼PCR检测引物进行河豚鱼成分的PCR检测与限制性内切酶NmeAⅢ酶切确证实验。18个供试样品中3个样品无PCR扩增产物,判为阴性结果,15个样品初步判为疑似阳性。应用限制性内切酶NmeAⅢ对疑似阳性样品的PCR产物进行酶切与电泳分析,电泳图谱与河豚鱼阳性对照不同的2个样品,判定为假阳性结果,电泳图谱与阳性对照相同的4个未知学名的河豚鱼加工样品,确证为阳性结果,即检出河豚鱼成分,PCR产物序列经GenBank同源性序列查询比对(BLAST)予以验证,建立了简便的河豚鱼成分PCR检测结果确证方法。False positive results frequently happen in PCR detection of animal and plant components.In order to exclude the possibility of false positive results,restriction endonuclease digestion was used to confirm the results of PCR for puffer fish components.Eighteen samples were detected,of which 3 samples were sentenced to be negative whereas the remaining 15 samples were suspected positive based on PCR results.Restriction endonuclease （NmeA Ⅲ） digestion and agarose gel electrophoresis were used to analyze the PCR products of the suspected positive puffer fish component.The electrophoresis pattems of PCR fragments digested by NmeA Ⅲ were different from the positive results of puffer fish in 2 non-puffer fish samples,which were judged as false positive.The electrophoresis patterns of 4 samples of processed puffer fish with unknown scientific names agreed with those of the puffer fish positive samples and were therefore confirmed.The PCR products were verified by GenBank DNA sequence homology sequence query （BLAST）.In conclusion,a simple method to confirm PCR results of puffer fish ingredients has been established in this study.

关 键 词：河豚鱼 PCR检测 限制性内切酶 DNA测序 结果确证 

分 类 号：S917.4[农业科学—水产科学] Q783.1[生物学—分子生物学]