| According to the target gene,specific primers were designed to explore the influence of various electric field factors on the virus content,and a suitable test electrode was designed to quantify the fitting effect of the equivalent circuit model.The physiological changes of tissue cells are studied and analyzed,and finally the cells are observed microscopically.The main work and conclusions of the thesis are as follows:(1)According to the nucleic acid information of apple necrosis mosaic virus in China published by NCBI,9 pairs of primers were designed by using Oligo 7.0 software through theoretical analysis of primer-dimer structure,primer hairpin structure and primer-template mismatch rate.Experimental verification selects an optimal primer for RT-QPCR.The results show that the high-voltage electric field can reduce the virus content,and the needle-shaped plate has the best effect among the three types of plates,and it is clear through the plate modeling and electric field simulation that the size of the plasma wind and the number of RONS species under the high-voltage electric field are responsible for the decrease in the virus content key reason.(2)By comparing the changes of impedance modulus value and phase angle under various voltage conditions,0.6 v is finally selected as the test voltage.The two curves are stable in the frequency range less than 1 MHz,and the final selection test range is 100 Hz?1 MHz.Three kinds of test electrodes were designed:acupuncture type,gel type and parallel plate type.The standard deviation of impedance mode value and phase angle of 5 repeated measurements at four frequency points were compared.Finally,the parallel plate type electrode was selected as the test electrode.Through multiple measurements of electrical impedance parameters on different parts of the same branch and the same part of different branches,it was found that there was no significant difference in impedance modulus and phase angle between groups.All can be used as processing objects of different processing groups.(3)The feasibility of the electrical impedance spectrum method was analyzed theoretically.According to the characteristics of the impedance spectrum,4 models were selected from the 6models to fit the impedance data.The spectrum data was substituted into different equivalent circuits,and complex numbers were used.The nonlinear least squares data processing method is used for theoretical analysis,and six indicators such as RMSE?SSE?R?R~2?DC and F-statistic are used to quantify the fitting effect.The results show that the modified Hayden model has the best fitting effect.(4)During the resting time,the impedance curve of the treatment group showed that it first decreased and then increased,followed by a short plateau and then continued to increase.When treated with an electric field for 150 min,the impedance curve showed a linear increase.The changes in the high frequency range of the impedance curve can be used to reflect the extreme damage of the electric field to the tissue.The Cole-cole chart of the treatment group before the treatment time of 90 min shows that the arc shrinks in a short time after leaving the electric field.The Cole-cole diagram of the treatment group with the treatment time of 120 min was close to linear change,which was related to the sharp increase of the phase angle.The circle graph at 150 min shows a linear change at the beginning,and then gradually shows a"scoop-shaped"change,which is related to the sharp drop of the impedance curve in the high frequency range.The higher the voltage or the longer the treatment time,the lower the cell membrane capacitance,and both the extracellular resistance and the intracellular resistance increased with the increase of resting time,applied voltage and treatment time.According to the observation by inverted fluorescence microscope,it was found that the phloem tissue cells of the treatment group contained large air bubbles,and the chloroplasts and other organelles were distributed around the cells. |
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