Development of an automated DNA purification module using a micro-fabricated pillar chip.
We present a fully automated DNA purification module comprised of a micro-fabricated chip and sequential injection analysis system that is designed for use within autonomous instruments that continuously monitor the environment for the presence of biological threat agents. The chip has an elliptical flow channel containing a bed (3.5 x 3.5 mm) of silica-coated pillars with height, width and center-to-center spacing of 200, 15, and 30 microm, respectively, which provides a relatively large surface area (ca. 3 cm(2)) for DNA capture in the presence of chaotropic agents. We have characterized the effect of various fluidic parameters on extraction performance, including sample input volume, capture flow rate, and elution volume. The flow-through design made the pillar chip completely reusable; carryover was eliminated by flushing lines with sodium hypochlorite and deionized water between assays. A mass balance was conducted to determine the fate of input DNA not recovered in the eluent. The device was capable of purifying and recovering Bacillus anthracis genomic DNA (input masses from 0.32 to 320 pg) from spiked environmental aerosol samples, for subsequent analysis using polymerase chain reaction-based assays.
Hindson
Benjamin J
BJ
Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
Gutierrez
Dora M
DM
Ness
Kevin D
KD
Makarewicz
Anthony J
AJ
Metz
Thomas R
TR
Setlur
Ujwal S
US
Benett
William B
WB
Loge
Jeffrey M
JM
Colston
Bill W
BW
Francis
Paul S
PS
Barnett
Neil W
NW
Dzenitis
John M
JM
D000336:Aerosols; D000393:Air Pollutants / Q000032:analysis*; D000818:Animals; D001408:Bacillus anthracis / Q000235:genetics*; D004269:DNA, Bacterial / Q000032:analysis*; D004784:Environmental Monitoring / Q000379:methods*; D004867:Equipment Design; D017022:Flow Injection Analysis; D006801:Humans; D008832:Microchemistry; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction / Q000379:methods
D000336:Aerosols; D000393:Air Pollutants; D004269:DNA, Bacterial