Abstract
BACKGROUND: Analysis of fungal genome sequence assemblies reveals that telomeres are poorly represented even though telomeric reads tend to be superabundant. We surmised that the problem might lie in the DNA shearing conditions used to create clone libraries for genome sequencing.
RESULTS: A shotgun strategy was used to sequence and assemble circular and linear cosmid DNAs sheared using conditions typical for a genome project. The DNA sheared in circular form assembled into a single sequence contig. However, the linearized cosmid produced an incomplete assembly because the two DNA termini, though greatly overrepresented in the clone library used for sequencing, were separated from neighboring sequences by gaps of approximately 1.4 and 1.8 kb. These gap sizes were reduced, but not eliminated, by shearing the linear cosmid into smaller fragments. Mapping of shearing breakpoints revealed a paucity of breaks in the subterminal regions of the linearized cosmid and also near chromosome ends of the fungus Neurospora crassa.
CONCLUSION: Together, our data indicate that the ends of linear DNA molecules are recalcitrant to hydrodynamic shearing. We propose that this causes DNA termini to be overrepresented in the resulting fragment population but ultimately prevents their incorporation into sequence assemblies.
Document Type
Article
Publication Date
2-2-2010
Digital Object Identifier (DOI)
http://dx.doi.org/10.1186/1471-2164-11-87
Repository Citation
Schwartz, Sherri L. and Farman, Mark L., "Systematic overrepresentation of DNA termini and underrepresentation of subterminal regions among sequencing templates prepared from hydrodynamically sheared linear DNA molecules" (2010). Plant Pathology Faculty Publications. 20.
https://uknowledge.uky.edu/plantpath_facpub/20
Additional file 1
Notes/Citation Information
Published in BMC Genomics, v. 11, 87.
© 2010 Schwartz and Farman; licensee BioMed Central Ltd.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.