Construction Of A PRV Mutant Strain SA215-T And Investigation Of Its Nerve Conduction Properties

Using Lipofectamine 3000 transfection reagent, plasmid of PP63 and pseudorabies virus Fa strain genomic DNA were transfected into BHK-21 cells, the recombinant virus gI/gE/Us9 deletion were screened by plaque, and named SA215-T.Using PCR, gene sequencing, western-blot, electron microscope, and the growth curve determination method for the detection of recombinant virus PRV SA215-T.The results show that:the recombinant virus SA215-T gI/gE/Us9 gene has effectively deleted, Western-blot was not stained in the absence of expression of gE gene, there is no obvious difference in the morphology. Growth curve of the recombinant virus and the parental virus in cells were both achievd a high viral titer.In order to explore three different virus strains-SA215-T, parent strain Fa, and Bartha-K61 on the nerve conduction function, this experiment with mice biceps reflex as a model, three kinds of strains were inoculated into mice biceps brachii. After the statistics, mice inoculated Fa with an average survival time of 73.33h, which was only half of the other two groups. And beginning at 60h, mice appeared obvious symptoms such as itching and anxiety, which were to blame for the toxicity effect of Fa. Acquisition of the dying mouse spinal cord and brain, using PCR method to detect pseudorabies virus is mainly distributed in the cervical spinal cord at C5-C6. Then established a method for detection of pseudorabies virus in spinal cord, nerve conduction study in mice, biceps, and the detection results are compared with those of PCR. The results showed, pseudorabies virus is mainly located in the cervical spinal cord at the positive staining of C5-C6, more gathered in the gray matter of the spinal cord motor neuron pool area (motorneuron pool), also found that some C5-C6 in the spinal cord were positive by the PCR test, while immunohistochemical detection is negative stained.