28 October 2009 Design of a real-time pyrosequencer by bioluminometric assay coupled with photodiode array
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Objective: A system for real-time pyrosequencing by biolumino-metric assay coupled with a photodiode array is presented. The approach is based on the detection of DNA polymerase activity by an enzymatic luminometric inorganic pyrophosphate (PPi) detection assay. It is designed to offer a convenient pyrosequencing-based platform for DNA sequencing with low cost and simple structure. Method: Together with the automated dNTPs micro-dispensers, a photoelectric conversion circuit with high sensitive photodiode S1133-01 and ultra-low bias current monolithic operational amplifier OPA129 is chosen to detect the luminometric, monitoring the ATP production continuously. The signal to noise ratio is improved efficiently by routing PCB and shielding carefully. Results: A C14 standard light source is selected to compare the sensitivity of the system to the photomultiplier tubes (PMT). The results show that the magnification of the circuits is ten times higher than the PMT, which is sufficient for detecting the DNAs as small as 10 fmol. Four different type of dNTPs are injected into the reaction cell in turn and the peak of luminescence illuminates the result, which is coinciding with the commercial system. Conclusion: The system is small, simple and inexpensive, which is enough for real-time pyrosequencing. But there are many works to do to construct the fast and high-flux DNA sequencing system.
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Haifeng Chen, Haifeng Chen, Chao Liang, Chao Liang, } "Design of a real-time pyrosequencer by bioluminometric assay coupled with photodiode array", Proc. SPIE 7519, Eighth International Conference on Photonics and Imaging in Biology and Medicine (PIBM 2009), 75190D (28 October 2009); doi: 10.1117/12.843545; https://doi.org/10.1117/12.843545

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