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Microarray generation of thousand-member oligonucleotide libraries.
PLoS One. 2011;6(9):e24906
Authors: Svensen N, Díaz-Mochón JJ, Bradley M
Abstract
The ability to efficiently and economically generate libraries of defined pieces of DNA would have a myriad of applications, not least in the area of defined or directed sequencing and synthetic biology, but also in applications associated with encoding and tagging. In this manuscript DNA microarrays were used to allow the linear amplification of immobilized DNA sequences from the array followed by PCR amplification. Arrays of increasing sophistication (1, 10, 3,875, 10,000 defined sequences) were used to validate the process, with sequences verified by selective hybridization to a complementary DNA microarray and DNA sequencing, which demonstrated a PCR error rate of 9.7×10(-3)/site/duplication. This technique offers an economical and efficient way of producing specific DNA libraries of hundreds to thousands of members with the DNA-arrays being used as "factories" allowing specific DNA oligonucleotide pools to be generated. We also found substantial variance observed between the sequence frequencies found via Solexa sequencing and microarray analysis, highlighting the care needed in the interpretation of profiling data.
PMID: 21966380 [PubMed - in process]
More...
Microarray generation of thousand-member oligonucleotide libraries.
PLoS One. 2011;6(9):e24906
Authors: Svensen N, Díaz-Mochón JJ, Bradley M
Abstract
The ability to efficiently and economically generate libraries of defined pieces of DNA would have a myriad of applications, not least in the area of defined or directed sequencing and synthetic biology, but also in applications associated with encoding and tagging. In this manuscript DNA microarrays were used to allow the linear amplification of immobilized DNA sequences from the array followed by PCR amplification. Arrays of increasing sophistication (1, 10, 3,875, 10,000 defined sequences) were used to validate the process, with sequences verified by selective hybridization to a complementary DNA microarray and DNA sequencing, which demonstrated a PCR error rate of 9.7×10(-3)/site/duplication. This technique offers an economical and efficient way of producing specific DNA libraries of hundreds to thousands of members with the DNA-arrays being used as "factories" allowing specific DNA oligonucleotide pools to be generated. We also found substantial variance observed between the sequence frequencies found via Solexa sequencing and microarray analysis, highlighting the care needed in the interpretation of profiling data.
PMID: 21966380 [PubMed - in process]
More...