title: A novel universal primer-multiplex-PCR method with sequencing gel electrophoresis analysis.
abstract: In this study, a novel universal primer-multiplex-PCR (UP-M-PCR) method adding a universal primer (UP) in the multiplex PCR reaction system was described. A universal adapter was designed in the 5'-end of each specific primer pairs which matched with the specific DNA sequences for each template and also used as the universal primer (UP). PCR products were analyzed on sequencing gel electrophoresis (SGE) which had the advantage of exhibiting extraordinary resolution. This method overcame the disadvantages rooted deeply in conventional multiplex PCR such as complex manipulation, lower sensitivity, self-inhibition and amplification disparity resulting from different primers, and it got a high specificity and had a low detection limit of 0.1 ng for single kind of crops when screening the presence of genetically modified (GM) crops in mixture samples. The novel developed multiplex PCR assay with sequencing gel electrophoresis analysis will be useful in many fields, such as verifying the GM status of a sample irrespective of the crop and GM trait and so on.
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China.
authors.forename: Wentao
authors.initials: W
authors.lastname: Xu
authors.forename: Zhifang
authors.initials: Z
authors.lastname: Zhai
authors.forename: Kunlun
authors.initials: K
authors.lastname: Huang
authors.forename: Nan
authors.initials: N
authors.lastname: Zhang
authors.forename: Yanfang
authors.initials: Y
authors.lastname: Yuan
authors.forename: Ying
authors.initials: Y
authors.lastname: Shang
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Luo
mesh_terms: D029688:Brassica napus / Q000235:genetics; D029441:Carica / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D018744:DNA, Plant / Q000032:analysis* / Q000235:genetics; D004586:Electrophoresis / Q000379:methods*; D003368:Gossypium / Q000235:genetics; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D012275:Oryza / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA; D013025:Glycine max / Q000235:genetics; D003313:Zea mays / Q000235:genetics
chemical_list: D017931:DNA Primers; D018744:DNA, Plant; D010940:Plant Proteins