shRNA capable of inhibiting replication and infection of Japanese encephalitis virus and application thereof
A technology of Japanese encephalitis virus and template sequence, applied in the field of genetic engineering
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Embodiment 1
[0034] The preparation of embodiment 1 shRNA
[0035] (1) Design and synthesis of oligonucleotides
[0036] Taking JEV's E, NS3, NS4b gene mRNA sequence as the target sequence ( figure 1 ), using Ambion’s siRNA design software, through homology analysis, designed and synthesized 4 for 1553~1571bp (E1), 1679~1697bp (E2), 6083~6101bp (NS3) and 7350~7368bp (NS4b) of the JEV gene, respectively. for shRNA oligonucleotide strands. TTCAAGAGA is selected for its loop structure to avoid the formation of termination signal, and the transcription termination sequence of shRNA adopts T6 structure. CACC is added to the 5' end of the sense strand template, which is complementary to the sticky end formed after digestion with BbsI; GATC is added to the 5' end of the antisense strand template, which is complementary to the sticky end formed after digestion with BamHI; if the target sequence of shRNA If the first base is not G, a G is added after the CACC of the template strand. The templa...
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