Linked Life Data

10217163

Source:http://linkedlifedata.com/resource/pubmed/id/10217163

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
1999-6-28
pubmed:abstractText
Reconstitution experiments were performed by using an ordinary dye-primer protocol of a template spiked with known amounts of truncated fragments. We observed that as little as 0.2 mole-percentage of the truncated fragment caused sequence interpretation problems. Two protocols were developed for sequencing with dye-labeled terminators; this eliminates the problems with truncated fragments, which are adapted to a one-dye chemistry. One was designed for single extension sequencing using T7 DNA polymerase and one for cycle sequencing. To avoid precipitation and centrifugation and to facilitate automation, the dye-terminator protocols included the use of a biotinylated sequencing primer. Thus, the Sanger fragments were recovered and, by magnetic separation, washed and released by formamide, EDTA, and heat treatment before loading on the electrophoresis gel. Integrated procedures for sequencing PCR products using one-dye-labeled terminators suitable for automation are described. High quality data in terms of long reads and detection of polymorphisms is obtained. The protocols serve as attractive alternatives to internal labeling and dye-primer approaches.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0173-0835
pubmed:author
pubmed:issnType
Print
pubmed:volume
20
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
502-10
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
1999
pubmed:articleTitle
Truncated fragments in polymerase chain reaction-based DNA sequencing.
pubmed:affiliation
Department of Biotechnology, Royal Institute of Technology, Stockholm, Sweden.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't