{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T07:45:35Z","timestamp":1792741535099,"version":"4.5.0"},"reference-count":31,"publisher":"National Academy of Sciences","issue":"7","content-domain":{"domain":["www.pnas.org"],"crossmark-restriction":true},"short-container-title":["Proc. Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[2004,2,17]]},"abstract":"<jats:p>\n                    Conventional asymmetric PCR is inefficient and difficult to optimize because limiting the concentration of one primer lowers its melting temperature below the reaction annealing temperature. Linear-After-The-Exponential (LATE)\u2013PCR describes a new paradigm for primer design that renders assays as efficient as symmetric PCR assays, regardless of primer ratio. LATE-PCR generates single-stranded products with predictable kinetics for many cycles beyond the exponential phase. LATE-PCR also introduces new probe design criteria that uncouple hybridization probe detection from primer annealing and extension, increase probe reliability, improve allele discrimination, and increase signal strength by 80\u2013250% relative to symmetric PCR. These improvements in PCR are particularly useful for real-time quantitative analysis of target numbers in small samples. LATE-PCR is adaptable to high throughput applications in fields such as clinical diagnostics, biodefense, forensics, and DNA sequencing. We showcase LATE-PCR via amplification of the cystic fibrosis\n                    <jats:italic>CF<\/jats:italic>\n                    \u0394\n                    <jats:italic>508<\/jats:italic>\n                    allele and the Tay-Sachs disease\n                    <jats:italic>TSD 1278<\/jats:italic>\n                    allele from single heterozygous cells.\n                  <\/jats:p>","DOI":"10.1073\/pnas.0305476101","type":"journal-article","created":{"date-parts":[[2004,2,17]],"date-time":"2004-02-17T15:32:02Z","timestamp":1077031922000},"page":"1933-1938","update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":193,"title":["Linear-After-The-Exponential (LATE)\u2013PCR: An advanced method of asymmetric PCR and its uses in quantitative real-time analysis"],"prefix":"10.1073","volume":"101","author":[{"given":"J. Aquiles","family":"Sanchez","sequence":"first","affiliation":[{"name":"Department of Biology, MS 008, Brandeis University, 415 South Street, Waltham, MA 02454-9110"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kenneth E.","family":"Pierce","sequence":"additional","affiliation":[{"name":"Department of Biology, MS 008, Brandeis University, 415 South Street, Waltham, MA 02454-9110"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"John E.","family":"Rice","sequence":"additional","affiliation":[{"name":"Department of Biology, MS 008, Brandeis University, 415 South Street, Waltham, MA 02454-9110"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lawrence J.","family":"Wangh","sequence":"additional","affiliation":[{"name":"Department of Biology, MS 008, Brandeis University, 415 South Street, Waltham, MA 02454-9110"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"341","published-online":{"date-parts":[[2004,2,9]]},"reference":[{"key":"e_1_3_2_1_2","doi-asserted-by":"publisher","DOI":"10.1007\/s000180050501"},{"key":"e_1_3_2_2_2","doi-asserted-by":"publisher","DOI":"10.1038\/335414a0"},{"key":"e_1_3_2_3_2","unstructured":"Kidd K. 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