Evaluation of methods for sequence analysis of highly repetitive DNA templates.
The DNA Sequencing Research Group (DSRG) of the ABRF conducted a study to assess the ability of DNA sequencing core facilities to successfully sequence a set of well-defined templates containing difficult repeats. The aim of this study was to determine whether repetitive templates could be sequenced accurately by using equipment and chemistries currently utilized in participating sequencing laboratories. The effects of primer and template concentrations, sequencing chemistries, additives, and instrument formats on the ability to successfully sequence repeat elements were examined. The first part of this study was an analysis of the results of 361 chromatograms from participants representing 40 different laboratories who attempted to sequence a panel of difficult-to-sequence templates using their best in-house protocols. The second part of this study was a smaller multi-laboratory evaluation of a single robust protocol with the same panel of templates. This study provides a measure of the potential success of different approaches to sequencing across homopolymer tracts and repetitive elements.
Hawes
John W
JW
Miami University, Oxford, OH, USA.
Knudtson
Kevin L
KL
Escobar
Helaman
H
Grills
George S
GS
Hunter
Timothy C
TC
Jackson-Machelski
Emily
E
Lin
Heather
H
Needleman
David S
DS
Pershad
Rashmi
R
Wiebe
Glenis J
GJ
D000818:Animals; D002845:Chromatography; D004247:DNA / Q000737:chemistry*; D017931:DNA Primers / Q000737:chemistry; D005069:Evaluation Studies as Topic; D051379:Mice; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D017421:Sequence Analysis / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods*
D017931:DNA Primers; D004247:DNA