Rapid recovery of DNA from agarose gel slices by coupling electroelution with monolithic SPE.
An amino silica monolithic column prepared by in situ polymerization of tetraethoxysilane and N-(beta-aminoethyl)-gamma-aminopropyltriethoxysilane was firstly applied to recover DNA from agarose gel slices by coupling electroelution with monolithic SPE. DNA was electroeluted from the agarose gel slices onto the amino silica monolithic column. The DNA adsorbed on this monolithic column was then recovered using sodium phosphate solution at pH 10. The whole recovery procedure could be completed within 10 min because the use of amino silica monolithic column accelerated the DNA capture and facilitated the DNA release. Electroelution conditions, such as buffer pH, buffer concentration and applied voltage, were online optimized. The average yield for herring sperm DNA, pBR 322 DNA and lambda DNA recovered from 1.0% w/v agarose gel slices were 55+/-4, 50+/-6 and 42+/-7% (n=3), respectively. The polymerase chain reaction performance of pGM plasmid recovered from agarose gel slices demonstrated that the method could provide high-quality DNA for downstream processes. The combination of electroelution with monolithic SPE allows a rapid, simple and efficient DNA recovery method. This technique is especially useful for applications that need to purify small starting amounts of DNA.
Yu
Shengbing
S
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, PR China.
Yang
Shuixian
S
Zhou
Ping
P
Zhou
Ke
K
Wang
Jing
J
Chen
Xiangdong
X
D000818:Animals; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D005399:Fishes; D006863:Hydrogen-Ion Concentration; D008297:Male; D012685:Sepharose / Q000737:chemistry*; D012821:Silanes / Q000737:chemistry; D012822:Silicon Dioxide / Q000737:chemistry; D012826:Silicone Elastomers / Q000737:chemistry; D052616:Solid Phase Extraction / Q000379:methods*; D013094:Spermatozoa / Q000737:chemistry
D012821:Silanes; D012826:Silicone Elastomers; C006194:aminoethyl-aminopropyl-trimethoxysilane; C040733:tetraethoxysilane; D012822:Silicon Dioxide; D004247:DNA; D012685:Sepharose