| Research purposes:This study simulates the change of concentration,temperature and other physical factors through experiments,effect on nucleic acid test results of SARS-COV-2 pseudo-virus,in order to grasp the influencing factors of nucleic acid detection process in clinical practice,it is used to guide the actual nucleic acid detection process,to achieve the purpose of making the test results more accurate.Research objects and methods:The specimen of this experiment was SARS-COV-2 pseudo-virus,source of specimens:positive control reagents in the 2019novel corona-virus ribonucleic acid(2019-nCoV RNA)liquid indoor quality control and2019-nCoV nucleic acid detection kit(fluorescent PCR method)centrally procured by Ben-Gang General Hospital of Liaoning Health Group.According to the experimental requirements,The specimens were subjected to different concentrations of dilution,warming at 56℃ for 30 min,and storage after exposure to air,etc,then the samples were extracted and amplified with nucleic acid,Summary,statistics and analysis of the obtained result data.Statistical analysis with SPSS19.0 software,measurement data conforming to normal distribution,expressed by mean±standard deviation((?)±s);Where the two groups of information were compared using independent samples t-test;Multi-group data comparison,using analysis of variance,among them,ANOVA test was used for homogeneous variance,and Welch test was used for non-homogeneous variance;The rates of the two groups were compared by the chi-square test for numerical data.All statistical calculations were bounded by P＜0.05(if P=0.000,expressed with P＜0.01),represents a statistically significant difference.Results:The difference of nucleic acid detection results of samples with different concentrations is statistically significant.The overall difference of CT value of ORFlab gene at different concentrations between the normal temperature group and the 56℃30min group was statistically significant(F=21027.74 and 37021.64,P＜0.01),after multiple comparisons,the difference between each group was statistically significant(P＜0.01);The overall difference of the CT value of N gene at different concentrations between the normal temperature group and the 56℃30min group was statistically significant(F=97473.80 and 111233.19,P＜0.01),after multiple comparisons,the difference between each group was statistically significant(P＜0.01).In terms of temperature influence,the difference in temperature on the detection rate of nucleic acid detection in specimens was statistically significant.The detection rate of nucleic acid in normal temperature group was 95.00%(57/60),the detection rate of nucleic acid detection in the 56℃30min group was 80.00%(48/60),the difference of positive rate between the two groups was statistically significant(χ~2=4.88,P＜0.05).There is no statistical difference between the nucleic acid detection results of high concentration samples before and after heating,however,low concentration samples have statistical significance for nucleic acid detection before and after heating.No statistically significant difference between the 1×,10×ORFlab gene and N gene normal temperature group and 56℃ 30 min heating group comparison(t=-0.431,-0.576,-0.165and-0.277,P=0.684,0.570,0.870 and 0.785,all P＞0.05);There were statistically significant differences between the 100×,1000×ORFlab genes and N genes in the normal temperature group and the 56℃ 30min heating group(t=-40.309,-149.959,-63.6915 and-86.417,all P＜0.01).There was no statistically significant difference in ORFlab gene between the conventional group,the group after 4 hours of air exposure,and the group kept at-20℃ for 8 hours,(F=0.005,P=0.995,P＞0.05),after multiple comparisons,there was no statistically significant difference in ORFlab gene between the groups(P＞0.05);There was no statistically significant difference in N gene between the conventional group,the4-hour group after air exposure and the 8-hour group stored at minus 20℃(F=1.274,P=0.293,P＞0.05),after multiple comparisons,there was no statistically significant difference in N gene among the groups(P＞0.05).Conclusion:2019 novel corona-virus nucleic acid detection results may be affected by sample concentration and heating inactivation,false negative results may appear in low concentration samples;The lower the concentration of the specimen,the greater the effect of heat inactivation on the results,even false negative results for weakly positive specimens,however,within a certain range,storage time has no effect on nucleic acid detection results. |
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