Linked Life Data

20693531

Source:http://linkedlifedata.com/resource/pubmed/id/20693531

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
19
pubmed:dateCreated
2010-10-28
pubmed:abstractText
To meet the growing demand for synthetic genes more robust, scalable and inexpensive gene assembly technologies must be developed. Here, we present a protocol for high-quality gene assembly directly from low-cost marginal-quality microarray-synthesized oligonucleotides. Significantly, we eliminated the time- and money-consuming oligonucleotide purification steps through the use of hybridization-based selection embedded in the assembly process. The protocol was tested on mixtures of up to 2000 oligonucleotides eluted directly from microarrays obtained from three different chip manufacturers. These mixtures containing <5% perfect oligos, and were used directly for assembly of 27 test genes of different sizes. Gene quality was assessed by sequencing, and their activity was tested in coupled in vitro transcription/translation reactions. Genes assembled from the microarray-eluted material using the new protocol matched the quality of the genes assembled from >95% pure column-synthesized oligonucleotides by the standard protocol. Both averaged only 2.7 errors/kb, and genes assembled from microarray-eluted material without clonal selection produced only 30% less protein than sequence-confirmed clones. This report represents the first demonstration of cost-efficient gene assembly from microarray-synthesized oligonucleotides. The overall cost of assembly by this method approaches 5¢ per base, making gene synthesis more affordable than traditional cloning.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
1362-4962
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
38
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
e180
pubmed:dateRevised
2011-8-24
pubmed:meshHeading
pubmed:year
2010
pubmed:articleTitle
High-quality gene assembly directly from unpurified mixtures of microarray-synthesized oligonucleotides.
pubmed:affiliation
Center for Innovations in Medicine of the Biodesign Institute at the Arizona State University, Tempe, AZ 85287, USA. aborovkov@synbuild.com
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Research Support, N.I.H., Extramural