Evaluation of Extraction Methodologies for Corn Kernel (Zea mays) DNA for Detection of Trace Amounts of Biotechnology-Derived DNA

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• 作者：Marcia J. Holden ; Joseph R. Blasic ; Jr. ; Louis Bussjaeger ; Chuan Kao ; Luke A. Shokere ; Donald C. Kendall ; Larry Freese ; and G. Ronald Jenkins
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2003
• 出版时间：April 23, 2003
• 年：2003
• 卷：51
• 期：9
• 页码：2468 - 2474
• 全文大小：80K
• 年卷期：v.51,no.9(April 23, 2003)
• ISSN：1520-5118

文摘

Sensitive and accurate testing for trace amounts of biotechnology-derived DNA from plant materialrequires pure, high-quality genomic DNA as template for subsequent amplification using thepolymerase chain reaction (PCR). Six methodologies were evaluated for extracting DNA from groundcorn kernels spiked with 0.1% (m/m) CBH351 (StarLink) corn. DNA preparations were evaluated forpurity and fragment size. Extraction efficiency was determined. The alcohol dehydrogenase gene(adh1) and the CBH351 (cry9C, 35S promoter) genes in the genomic DNA were detected usingPCR. DNA isolated by two of the methods proved unsuitable for performing PCR amplification. Allother methods produced some DNA preparations that gave false negative PCR results. We observedthat cornstarch, a primary component of corn kernels, was not an inhibitor of PCR, while acidicpolysaccharides were. Our data suggest that amplification of an endogenous positive control gene,as an indicator for the absence of PCR inhibitors, is not always valid. This study points out aspectsof DNA isolation that need to be considered when choosing a method for a particular plant/tissuetype.Keywords: Zea mays; corn; genomic; DNA; extraction; polymerase chain reaction; PCR; PCR inhibition