Journal
NUCLEIC ACIDS RESEARCH
Volume 40, Issue 11, Pages -Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gks230
Keywords
-
Categories
Funding
- National Institutes of Health [R01AI076247]
- National Science Foundation's Frontiers in Integrative Biological Research [FIBR-0527023]
- NSF's BEACON Center for the Study of Evolution in Action [DBI-0939454]
Ask authors/readers for more resources
Isothermal nucleic acid amplification is becoming increasingly important for molecular diagnostics. Therefore, new computational tools are needed to facilitate assay design. In the isothermal EXPonential Amplification Reaction (EXPAR), template sequences with similar thermodynamic characteristics perform very differently. To understand what causes this variability, we characterized the performance of 384 template sequences, and used this data to develop two computational methods to predict EXPAR template performance based on sequence: a position weight matrix approach with support vector machine classifier, and RELIEF attribute evaluation with Naive Bayes classification. The methods identified well and poorly performing EXPAR templates with 67-70% sensitivity and 77-80% specificity. We combined these methods into a computational tool that can accelerate new assay design by ruling out likely poor performers. Furthermore, our data suggest that variability in template performance is linked to specific sequence motifs. Cytidine, a pyrimidine base, is over-represented in certain positions of well-performing templates. Guanosine and adenosine, both purine bases, are over-represented in similar regions of poorly performing templates, frequently as GA or AG dimers. Since polymerases have a higher affinity for purine oligonucleotides, polymerase binding to GA-rich regions of a single-stranded DNA template may promote non-specific amplification in EXPAR and other nucleic acid amplification reactions.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available