Fluorogenic ribonuclease protection (FRIP) analysis of single nucleotide polymorphisms (SNPs) in Japanese Rice (Oryza sativa L.) DNA for cultivar discrimination.
A rapid and easy method to discriminate plant cultivars is indispensable to confirm food labeling. We established a fluorogenic ribonuclease protection (FRIP) assay to discriminate Japanese rice (Oryza sativa L.) cultivars based on single nucleotide polymorphisms (SNPs). The FRIP assay uses a hybridization technique between fluorescent probes and the target sequence prepared by run-off transcription, but the requirement of two PCR thermocycles is the problem when preparing template DNA for run-off transcription from rice genomic DNA. In this study, we designed new PCR primers with asymmetric melting temperatures. These primers amplified the target SNP marker containing a T7 RNA polymerase promoter sequence upstream of the target sequence in a single PCR. Moreover, 100 cultivars were discriminated with the patterns of 15 SNPs. The assay can be used as a rapid method of analysis to discriminate Japanese rice cultivars.
Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, Fukuoka, Japan.
Momoko
M
Kitaoka
Takuya
T
Wada
Takeshi
T
Nishio
Masahiro
M
Goto
D004252:DNA Mutational Analysis; D018744:DNA, Plant / Q000032:analysis; D005515:Food Labeling; D018745:Genome, Plant; D012275:Oryza / Q000235:genetics*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D013045:Species Specificity
D018744:DNA, Plant