Extraction of genomic DNA using a new amino silica monolithic column.
A new amino silica monolithic column was developed for DNA extraction in a miniaturized format. The monolithic column was prepared in situ by polymerization of tetraethoxysilane (TEOS) and N-(beta-aminoethyl)-gamma-aminopropylmethyldimethoxysilane (AEAPMDMS). DNA was loaded in 50 mM tris(hydroxylmethyl)aminomethane-EDTA buffer at pH 7.0 and eluted with 300 mM potassium phosphate solution at pH 10.0. Under optimal condition, a 6.0-cm monolithic column provided a capacity of 56 ng DNA with an extraction efficiency of 71 +/- 5.2% (X +/- RSD). When the amino silica monolithic column was applied to extract genomic DNA from the whole blood of crucian carp, an extraction efficiency of 52 +/- 5.6% (X +/- RSD) was obtained by three extractions. Since the chaotropic-based sample loading and organic solvent wash steps were avoided in this procedure, the purified DNA was suitable for downstream processes such as PCR. This amino silica monolithic column was demonstrated to allow rapid and efficient DNA purification in microscale.
Liu
Lijia
L
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, China.
Yu
Shengbing
S
Yang
Shuixian
S
Zhou
Ping
P
Hu
Jiming
J
Zhang
Yibing
Y
D000199:Actins / Q000235:genetics; D000818:Animals; D002347:Carps / Q000235:genetics; D002853:Chromatography, Liquid / Q000295:instrumentation* / Q000379:methods; D004247:DNA / Q000302:isolation & purification*; D016678:Genome; D044085:Microfluidics; D008855:Microscopy, Electron, Scanning; D012822:Silicon Dioxide / Q000737:chemistry*
D000199:Actins; D012822:Silicon Dioxide; D004247:DNA