Dextran sulfate provides a quantitative and quick microarray hybridization reaction.
Microarray technology is a powerful tool to speed up genomics study, yet many technical aspects need to be improved. The hybridization reaction of microarray experiments is carried out for 16h or overnight in order to obtain reasonably strong signals for analysis in the presence of high salt buffer, like SSC. However, the quantitative aspect of microarray hybridization has seldom been investigated. In this study, we showed that higher overall signals from hybridization were achieved in a buffer system containing dextran sulfate, which can accelerate the kinetics of reaction by increasing the local concentration of the reactants. The dextran sulfate containing hybridization solution increases the reaction 4-fold (median) for cDNA microarray and 29-fold for oligonucleotide microarray. More importantly, the solution also provides a quantitative hybridization reaction, where the hybridization signals are proportional to the abundance of transcript added. The enhancement in the kinetics of hybridization is due to both dextran sulfate and formamide present in the solution, but the effect is not due to the higher temperature used during the reaction. With a slightly longer reaction time the hybridization reaction with the solution allows the detection of hybridization signals from rare transcripts that is not possible with regular hybridization buffers. With appropriate washing, the enhancement of kinetics by the solution does not increase the background signals at all, allowing higher signal-to-noise ratios to be achieved.
Ku
Wei-Chi
WC
Department of Research and Development, U-Vision Biotech, Inc., Taipei, Taiwan, ROC.
Lau
Wai Kwan
WK
Tseng
Yu-Tien
YT
Tzeng
Chi-Meng
CM
Chiu
Sung-Kay
SK
D017360:Arabidopsis / Q000235:genetics; D002021:Buffers; D002232:Carbocyanines / Q000737:chemistry; D002460:Cell Line; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D016264:Dextran Sulfate / Q000737:chemistry*; D005559:Formamides / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D009693:Nucleic Acid Hybridization / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis; D012680:Sensitivity and Specificity
D002021:Buffers; D002232:Carbocyanines; D018076:DNA, Complementary; D005559:Formamides; D012333:RNA, Messenger; C089187:cyanine dye 3; C085321:cyanine dye 5; C031066:formamide; D016264:Dextran Sulfate