{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,10]],"date-time":"2026-11-10T12:33:24Z","timestamp":1794314004404,"version":"4.9.0"},"reference-count":28,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2020,7,23]],"date-time":"2020-07-23T00:00:00Z","timestamp":1595462400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2020,7,23]],"date-time":"2020-07-23T00:00:00Z","timestamp":1595462400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100012457","name":"Chongqing Basic and Frontier Research Project","doi-asserted-by":"publisher","award":["CSTC2018jcyjAX0218"],"award-info":[{"award-number":["CSTC2018jcyjAX0218"]}],"id":[{"id":"10.13039\/501100012457","id-type":"DOI","asserted-by":"publisher"}]},{"name":"the translational medical project of Children's Hospital of Chongqing Medical University","award":["Lcyj2015-1"],"award-info":[{"award-number":["Lcyj2015-1"]}]},{"name":">Chongqing Yuzhong District Sci"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Leber hereditary optic neuropathy (LHON) is a degenerative disease of the optic nerve associated with one of three mitochondrial DNA (mtDNA) m.3460G&gt;A, m.11778G&gt;A and m.14484T&gt;C mutations. Although several procedures are available to genotype these mutations, quantitative approaches with rapid, low-cost and easy to handle advantages for three LHON mtDNA mutations are rarely reported. Here, we firstly developed a \u201cone-step\u201d tetra-primer amplification-refractory mutation system (T-ARMS) PCR for qualitative genotyping of three LHON mtDNA mutations. Subsequently, we established single, duplex and triplex TaqMan MGB probe-based fluorescence quantitative PCR (qPCR) assays to perform both qualitative and quantitative analyses of three LHON mtDNA mutations. Standard curves based on tenfold diluted plasmid standard exhibited high specificity and sensitivity, stable repeatability and reliable detectable ability of TaqMan probe qPCR assays without cross-reactivity upon probes combination. Moreover, by comparing with SYBR Green qPCR, we further validated the feasibility of the triplex-probe qPCR assay for the quantitative detection of mtDNA copy number in blood samples. In conclusion, our study describes a rapid, low-cost, easy to-handle, and high-throughput TaqMan-MGB probe qPCR assay to perform both qualitative and quantitative analysis of three primary LHON mtDNA mutations, offering a promising approach for genetic screening and testing of LHON mutations.<\/jats:p>","DOI":"10.1038\/s41598-020-69220-7","type":"journal-article","created":{"date-parts":[[2020,7,23]],"date-time":"2020-07-23T06:04:12Z","timestamp":1595484252000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":22,"title":["TaqMan-MGB probe quantitative PCR assays to genotype and quantify three mtDNA mutations of Leber hereditary optic neuropathy"],"prefix":"10.1038","volume":"10","author":[{"given":"Bingqian","family":"Xue","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yang","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xin","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rui","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xin","family":"Zeng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Meihua","family":"Yang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaohui","family":"Xu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tingbo","family":"Ye","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liming","family":"Bao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yi","family":"Huang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2020,7,23]]},"reference":[{"key":"69220_CR1","doi-asserted-by":"publisher","first-page":"1427","DOI":"10.1126\/science.3201231","volume":"242","author":"DC Wallace","year":"1988","unstructured":"Wallace, D. C. et al. Mitochondrial DNA mutation associated with Leber\u2019s hereditary optic neuropathy. Science 242, 1427\u20131430. https:\/\/doi.org\/10.1126\/science.3201231 (1988).","journal-title":"Science"},{"key":"69220_CR2","doi-asserted-by":"publisher","first-page":"214","DOI":"10.1038\/883","volume":"19","author":"S DiMauro","year":"1998","unstructured":"DiMauro, S. & Schon, E. A. Nuclear power and mitochondrial disease. Nat. Genet. 19, 214\u2013215. https:\/\/doi.org\/10.1038\/883 (1998).","journal-title":"Nat. Genet."},{"key":"69220_CR3","doi-asserted-by":"publisher","first-page":"610","DOI":"10.1038\/nature08802","volume":"464","author":"Y He","year":"2010","unstructured":"He, Y. et al. Heteroplasmic mitochondrial DNA mutations in normal and tumor cells. Nature 464, 610\u2013614. https:\/\/doi.org\/10.1038\/nature08802 (2010).","journal-title":"Nature"},{"key":"69220_CR4","doi-asserted-by":"publisher","first-page":"638","DOI":"10.1056\/NEJM199509073331007","volume":"333","author":"DR Johns","year":"1995","unstructured":"Johns, D. R. Seminars in medicine of the Beth Israel Hospital, Boston. Mitochondrial DNA and disease. N. Engl. J. Med. 333, 638\u2013644. https:\/\/doi.org\/10.1056\/NEJM199509073331007 (1995).","journal-title":"N. Engl. J. Med."},{"key":"69220_CR5","doi-asserted-by":"publisher","first-page":"2362","DOI":"10.1167\/iovs.06-0331","volume":"48","author":"DF Ventura","year":"2007","unstructured":"Ventura, D. F. et al. Male prevalence of acquired color vision defects in asymptomatic carriers of Leber\u2019s hereditary optic neuropathy. Invest. Ophthalmol. Vis. Sci. 48, 2362\u20132370. https:\/\/doi.org\/10.1167\/iovs.06-0331 (2007).","journal-title":"Invest. Ophthalmol. Vis. 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Detection of mitochondrial DNA mutations using temporal temperature gradient gel electrophoresis. Electrophoresis 25, 2602\u20132610. https:\/\/doi.org\/10.1002\/elps.200406016 (2004).","journal-title":"Electrophoresis"},{"key":"69220_CR9","doi-asserted-by":"publisher","first-page":"432","DOI":"10.1086\/302996","volume":"67","author":"LA Tully","year":"2000","unstructured":"Tully, L. A. et al. A sensitive denaturing gradient-Gel electrophoresis assay reveals a high frequency of heteroplasmy in hypervariable region 1 of the human mtDNA control region. Am. J. Hum. Genet. 67, 432\u2013443. https:\/\/doi.org\/10.1086\/302996 (2000).","journal-title":"Am. J. Hum. Genet."},{"key":"69220_CR10","doi-asserted-by":"publisher","first-page":"1390","DOI":"10.1002\/mus.880181208","volume":"18","author":"Y Tanno","year":"1995","unstructured":"Tanno, Y. et al. Quantitation of heteroplasmy of mitochondrial tRNA(Leu(UUR)) gene using PCR-SSCP. 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