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Nested Machine Learning Facilitates Increased Sequence Content for Large-Scale Automated High Resolution Melt Genotyping.

Abstract：

:High Resolution Melt (HRM) is a versatile and rapid post-PCR DNA analysis technique primarily used to differentiate sequence variants among only a few short amplicons. We recently developed a one-vs-one support vector machine algorithm (OVO SVM) that enables the use of HRM for identifying numerous short amplicon sequences automatically and reliably. Herein, we set out to maximize the discriminating power of HRM + SVM for a single genetic locus by testing longer amplicons harboring significantly more sequence information. Using universal primers that amplify the hypervariable bacterial 16 S rRNA gene as a model system, we found that long amplicons yield more complex HRM curve shapes. We developed a novel nested OVO SVM approach to take advantage of this feature and achieved 100% accuracy in the identification of 37 clinically relevant bacteria in Leave-One-Out-Cross-Validation. A subset of organisms were independently tested. Those from pure culture were identified with high accuracy, while those tested directly from clinical blood bottles displayed more technical variability and reduced accuracy. Our findings demonstrate that long sequences can be accurately and automatically profiled by HRM with a novel nested SVM approach and suggest that clinical sample testing is feasible with further optimization.

journal_name

Sci Rep

journal_title

Scientific reports

authors

Fraley SI,Athamanolap P,Masek BJ,Hardick J,Carroll KC,Hsieh YH,Rothman RE,Gaydos CA,Wang TH,Yang S

doi

10.1038/srep19218

subject

Has Abstract

pub_date

2016-01-18 00:00:00

pages

19218

issn

2045-2322

pii

srep19218

journal_volume

6

pub_type

杂志文章

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