Abstract
Polymerase chain reaction (PCR) has become the mainstay of DNA sequence analysis. Yet there is always uncertainty concerning the source of the template DNA that gave rise to a particular PCR product. The risks of contamination, biased amplification, and product redundancy are especially high when limited amounts of template DNA are used. We have developed and applied molecular encoding principles to solve this source-uncertainty problem for DNA sequences generated by standard PCR. Batch-stamps specify the date and sample identity, and barcodes detect template redundancy. Our approach thus enables classification of each PCR-derived sequence as valid, contaminant, or redundant, and provides a measure of sequence diversity. We recommend that batch-stamps and barcodes be used when amplifying irreplaceable DNAs and cDNAs available for forensic, clinical, single cell, and ancient DNA analyses.
Similar content being viewed by others
References
Cohn DH, Starman BJ, Blumberg B, Byers PH (1990) Recurrence of lethal osteogenesis imperfecta due to parental mosaicism for a dominant mutation in a human type I collagen gene (COL1A1). Am J Hum Genet 46(3):591–601
Kolata G (2007) Faith in quick test leads to epidemic that wasn’t. In: New York Times, 22 Jan 2007
Laird CD, Pleasant ND, Clark AD, Sneeden JL, Hassan KM, Manley NC, Vary JC Jr., Morgan T, Hansen RS, Stöger R (2004) Hairpin-bisulfite PCR: assessing epigenetic methylation patterns on complementary strands of individual DNA molecules. Proc Natl Acad Sci USA 101(1):204–209
Lewontin RC (1994) The use of DNA profiles in forensic contexts. Stat Sci 9(2):259–262
Li HH, Gyllensten UB, Cui XF, Saiki RK, Erlich HA, Arnheim N (1988) Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature 335(6189):414–417
Miner BE, Stöger R, Burden AF, Laird CD, Hansen RS (2004) Molecular barcodes detect redundancy and contamination in hairpin-bisulfite PCR. Nucleic Acids Res 32(17):e135
Stöger R, Kajimura TM, Brown WT, Laird CD (1997) Epigenetic variation illustrated by DNA methylation patterns of the fragile-X gene FMR1. Hum Mol Genet 6(11):1791–1801
Wong DJ, Barrett MT, Stöger R, Emond MJ, Reid BJ (1997) p16INK4a promoter is hypermethylated at a high frequency in esophageal adenocarcinomas. Cancer Res 57(13):2619–2622
Acknowledgments
This work was supported by the National Institutes of Health grants GM 53805, HD 02274, and by the Washington Research Foundation. The method presented here is included in United States Patent Application 20070020640, filed by the University of Washington. We thank Carl Bergstrom, Alice Burden, Diane Genereux, Brooks Miner, and Jessica Sneeden for helpful suggestions and discussion.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
McCloskey, M.L., Stöger, R., Hansen, R.S. et al. Encoding PCR Products with Batch-stamps and Barcodes. Biochem Genet 45, 761–767 (2007). https://doi.org/10.1007/s10528-007-9114-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10528-007-9114-x