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Screening and construct-specific detection methods of transgenic Huafan No 1 tomato by conventional and real-time PCR

Author:
Yang, L., Shen, H., Pan, A., Chen, J., Huang, C., Zhang, D.
Source:
Journal of the science of food and agriculture 2005 v.85 no.13 pp. 2159-2166
ISSN:
0022-5142
Subject:
Solanum lycopersicum var. lycopersicum, tomatoes, transgenic plants, transgenes, antisense DNA, Cauliflower mosaic virus, promoter regions, polymerase chain reaction, DNA primers, nucleotide sequences, genetically modified foods, food analysis, 1-aminocyclopropane-1-carboxylate synthase
Abstract:
Genetically modified (GM) tomatoes have been approved for commercialization in many countries since the first GM tomato FLAVR SAVR was permitted for planting in 1994. In China, GM tomato Huafan No 1 with a character of long shelf-life was the first GM plant which was approved for commercialization in 1996. To meet the requirement of the GM tomatoes labeling policy that has been actualized in China since 2001, screening and construct-specific PCR detection methods for detecting the universal elements transformed into tomato, such as cauliflower mosaic virus 35S (CaMV35s) promoter and the nopaline synthase (NOS) terminator of Agrobacterium tumefaciens, and the specifically inserted heterologous DNA sequence between CaMV35s promoter and anti-sense ethylene-forming enzyme (EFE) gene were set up. To make the detection methods normative, a novel single copy tomato gene LAT52 was also used as an endogenous reference gene in the PCR detection systems. The limit of detection of screening and construct specific detection methods for Huafan No 1 was 68 haploid genome copies in conventional PCR detection, and three copies in TaqMan real-time PCR detection. The limit of quantitation of screening quantitative PCR assays for Huafan No 1 was three copies and was 25 copies for construct-specific quantitative PCR. Two samples with known Huafan No 1 tomato content were detected using the established conventional and real-time PCR systems, and these results also indicated that the established Huafan No 1 screening and construct-specific PCR detection systems were reliable, sensitive and accurate.
Agid:
1398872