title: Identification of WA-type three-line hybrid rice with real-time polymerase chain reaction (PCR) method.
abstract: A real-time fluorescent PCR (RTF-PCR) was developed to detect and quantify wild abortive (WA)-type three-line hybrid rice (Oryza sativa L.). The mitochondrial R₂₋₆₃₀ WA gene was reported to be closely related to male sterility in plants, and developed as a molecular maker to identify the cytoplasmic male sterility system of hybrid rice. First, we got the DNA sequence of R₂₋₆₃₀ WA gene in 17 rice species with traditional PCR. Then, a pair of specific primers (P₃, P₄) and TaqMan fluorescence probe (P₃₋₁₄) were designed based on the R₂₋₆₃₀ DNA sequence. The following RTF-PCR was performed on the 17 rice species finally. The results indicate that the probes used here are specific for three-line hybrid rice F₁ and male sterile lines. We can even identify a single hybrid seed using the probes, which confirmed that the probes can be applied to the identification and quantification of the WA-type three-line hybrid rice. In addition, the RFT-PCR system can be optimized when the annealing temperature is 60 °C and the Mg²⁺ concentration is 3.5 mmol/L.
authors.affiliation: Institute of Bast-Fiber crops, Chinese Academy of Agricultural Sciences, Xianjiahu West Road 348, Changsha, China. charles_chengyi@yahoo.com.cn
authors.forename: Y
authors.initials: Y
authors.lastname: Cheng
authors.forename: B D
authors.initials: BD
authors.lastname: Gao
authors.forename: H Y
authors.initials: HY
authors.lastname: Chen
authors.forename: J J
authors.initials: JJ
authors.lastname: Mao
authors.forename: A X
authors.initials: AX
authors.lastname: Cao
authors.forename: J G
authors.initials: JG
authors.lastname: Zhu
authors.forename: S F
authors.initials: SF
authors.lastname: Zhu
mesh_terms: D001483:Base Sequence; D002678:Chimera / Q000235:genetics; D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D004272:DNA, Mitochondrial / Q000145:classification / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics*; D005453:Fluorescence; D017343:Genes, Plant; D005838:Genotype; D008969:Molecular Sequence Data; D058889:Molecular Typing; D012275:Oryza / Q000145:classification / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012098:Reproduction
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial; D018744:DNA, Plant