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Objective To develop a multiplex real -time RT -PCR assay for simultaneous detection of enteroviruses and differentiation of EV71 and CA16.Methods Specific primers and probes were designed for enteroviruses,EV71 and CA16.The probes labeled with various fluorescent reporter dyes,and a triplex real -time RT -PCR technique was developed to simultaneously detect these viruses.A total of 91 clinical specimens with suspected HFMD were analyzed by this method.Results This assay could simultaneously detect enterovirus and differentiation of EV71 and CA16,and the sensitivity of the assay was up to 0.1 TCID50 /mL,and only need 2 to 3 hours for completing the detection.A total of 91 clinical specimens were detected by this assay in 28 of the 91(30.77%)specimens contained EV71,9 of the 91(9.89%) contained CA16,and 5 of the 91 (5.49%)contained other enteroviruses.Conclusion This assay would be a useful molecular diagnostic tool for large -scale screening of clinical samples,especially at the peroid of HFMD outbreaks.
ABSTRACT
0.05).Chlorpyrifos increased the activity of CYP1A1 in lungs,CYP2B1 in kidneys and lungs.Cypermethrin increased activity of CYP1A1 in livers,kidneys and lungs,CYP2B1 in livers and lungs,CYP3A in livers.These increases were statistically significant (P
ABSTRACT
Based on the conserved region nucleotide sequences of five potato viruses/viroid(Alfalfa Mosaic Virus,ALMV;Cucumber mosaic virus,CMV;Cucumber mosaic virus-satellite,CMV-sat;Potato virus Y,PVY;Potato spindle tuber viroid,PSTVd) and one inner control(18S rRNA),the microarray containing specific oligonucleotide probes and PCR probes were designed and fabricated.The effects of probe concentration,hybridization time,hybridization temperature and spotting solutions on microarray hybridiazation were evaluated.Finally the specificity of optimized plant virus detection array was validated.No significant effect on hybridization signal intensity was observed when the concentration of the oligonucleotide probes ranged from 5 to 20 ?mol/L,there was a linear relationship between the concentration of PCR probe and hybridization signal intensity.The greatest signal intensity were obtained when hybridized at 45℃ for 4 h,and the oligonucleotide probes and PCR probes had a similar effect on microarray hybrization.Among the different spotting solutions,DMSO produced a good reproducibility.The plant virus could be detected specifically by oligonucleotide probe microarray and PCR probe microarray after optimization.