Vortex fluidics-mediated DNA rescue from formalin-fixed museum specimens.

Autor: Totoiu CA; Department of Chemistry, University of California, Irvine, California, United States of America., Phillips JM; Flinders Institute for Nanoscale Science and Technology, College of Science and Engineering, Flinders University, Adelaide, South Australia, Australia., Reese AT; Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, Massachusetts, United States of America., Majumdar S; Department of Chemistry, University of California, Irvine, California, United States of America., Girguis PR; Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, Massachusetts, United States of America., Raston CL; Flinders Institute for Nanoscale Science and Technology, College of Science and Engineering, Flinders University, Adelaide, South Australia, Australia., Weiss GA; Department of Chemistry, University of California, Irvine, California, United States of America.; Department of Molecular Biology and Biochemistry, University of California, Irvine, California, United States of America.; Department of Pharmaceutical Sciences, University of California, Irvine, California, United States of America.
Jazyk: angličtina
Zdroj: PloS one [PLoS One] 2020 Jan 30; Vol. 15 (1), pp. e0225807. Date of Electronic Publication: 2020 Jan 30 (Print Publication: 2020).
DOI: 10.1371/journal.pone.0225807
Abstrakt: DNA from formalin-preserved tissue could unlock a vast repository of genetic information stored in museums worldwide. However, formaldehyde crosslinks proteins and DNA, and prevents ready amplification and DNA sequencing. Formaldehyde acylation also fragments the DNA. Treatment with proteinase K proteolyzes crosslinked proteins to rescue the DNA, though the process is quite slow. To reduce processing time and improve rescue efficiency, we applied the mechanical energy of a vortex fluidic device (VFD) to drive the catalytic activity of proteinase K and recover DNA from American lobster tissue (Homarus americanus) fixed in 3.7% formalin for >1-year. A scan of VFD rotational speeds identified the optimal rotational speed for recovery of PCR-amplifiable DNA and while 500+ base pairs were sequenced, shorter read lengths were more consistently obtained. This VFD-based method also effectively recovered DNA from formalin-preserved samples. The results provide a roadmap for exploring DNA from millions of historical and even extinct species.
Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: GAW and CLR, through their universities, have filed a patent titled "Method for Improving Protein Functionality using Vortexing Fluid Shear Forces (U.S. App No. 14/913,951). Debut Biotechnology, a company co-founded by GAW, has licensed this patent. GAW and CLR both serve on the Scientific Advisory Board of Debut Biotechnology and own shares of stock in the company. This does not alter our adherence to PLOS ONE policies on sharing data and materials.
Databáze: MEDLINE
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