Development of quality control and evaluation techniques for dried sheets of Asakusa-nori: Simple estimation of <i>Pyropia tenera</i> thallus content in dried sheets using PCR analysis

Abstract

In order to develop quality control and evaluation techniques for dried sheets of Asakusa-nori, a simple, rapid method was created for using PCR analysis to estimate Pyropia tenera thallus content. Based on the mitochondrial DNA (mtDNA) nucleotide sequences of P. tenera and P. yezoensis, PCR primer sets were designed to amplify two mtDNA regions (orf169 trnV and cox1 regions) with nucleotide polymorphisms between both species. After PCR ampliˆcation of P. tenera and P. yezoensis total DNAs with the mtDNA-speciˆc primers, structural diŠerences between both species such as gene loss and rearrangement in the orf169 trnV and cox1 regions were easily detected, indicating that PCR analysis of these mtDNA regions is useful for discriminating P. tenera from P. yezoensis. Pyropia tenera content consisting of thallus pieces randomly collected from commercially-available sheets of dried Asakusa-nori was successfully estimated by PCR analysis of these mtDNA regions. In addition, the mtDNA fragments originating from P. tenera and P. yezoensis were ampliˆed by competitive-PCR using total DNAs extracted from the punches of the dried-Asakusa nori sheets as the template. These results indicate that these techniques enable rapid, simple estimation of P. tenera thallus content in dried-Asakusa nori sheets.