Some U.S bioethanol plants use a blend of maize and sorghum grains as feedstocks depending on availability and price. Knowledge of the grain composition of milled feedstock is beneficial for maximizing process efficiency and for potential future regulatory requirements. To analyze feedstock composition, we developed a duplex real-time PCR assay based on TaqMan™ probes for the simultaneous analysis of flour blends. For sorghum, the method amplified a sequence within the F-Box domain-containing protein and for maize, the method amplified a sequence of the high mobility group a (HMGa) proteins. To acquire fluorescent signals simultaneously, FAM and TAMRA fluorescent probes were used to label the sorghum- and maize- specific probes, respectively. Results showed that the primers and probes were highly specific either for maize or sorghum. The real-time duplex PCR assay was validated on simulated flour binary mixtures and high in the precision and accuracy for the sorghum PCR system. Results with the established method showed specificity and sensitivity in quantifying sorghum composition in bioethanol feedstock mixtures. Resources in this dataset: Resource title: Primer sets used File name: Table 1 1.jpg Resource title: Experimental setup File name: experiemntal set ups Real time PCR project Sorgh-Maize.xlsx Resource title: Multiplex data HMGa and FBox File name: 2022-01-24 multiplex FBox - HMGa 60 C 200nM.xls Resource description: Data and analysis of sorhum maize and miusxtures using HMGa and FBox genes Resource title: multiplex FBox HMGa flour mixes 60 C File name: 2022-02-22 multiplex FBox HMGa flour mixes 60 C 11 points.xls Resource description: PCR multiplex data FBox HMGa flour mixes 60 C Resource title: Standard curves for zein DNA HMGa gene File name: 2022-01-11 Hmga - Zein X07535 standard curves.xls Resource description: Standard curves for zein DNA HMGa gene detection

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  Primer sets used
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  Experimental setup
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  Multiplex data HMGa and FBox
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  multiplex FBox HMGa flour mixes 60 C
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  Standard curves for zein DNA HMGa gene
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