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[Objective ] pokeweed antiviral protein (pokeweed antiviral proteins, PAPs) is a synthesis of different organizations or different growth stages pokeweed (phytolacca amercana) , a class of enzyme function ribosome inactivating protein ( ribosome - inactivating proteins , RIPs), there are a variety of types , PAP- I , PAP - II , PAP- III , PAP -S , PAP- R from the leaves of spring , early summer leaves , late summer leaves , seeds, roots . Pokeweed antiviral protein - Ⅱ (pokeweed antiviral protein Ⅱ), with RNA N- glycosidase activity, specificity to 4324 of the 28S rRNA from eukaryotic ribosomes and prokaryotic ribosomal 23S rRNA 2600 cutting the next adenine, interference extension factor EF-2 catalyst of GTP hydrolysis , thereby preventing protein synthesis . Recent research has shown that the PAP-Ⅱ protein can inhibit a variety of viruses , including the HIV virus , research will construct immunotoxins for killing tumor cells . The laboratory using genetic engineering methods , the PAP-Ⅱ gene was successfully cloned from the fresh leaves of pokeweed summer , and inserted into the prokaryotic expression vector , and successfully expressed in E. coli BL21 PAP-Ⅱ protein , as further study of the PAP-Ⅱ anti-viral , anti- cancer mechanism to provide a research platform . [ Method ] reported cDNA sequence , designed and synthesized two primers , two primers were added to the Hind III and Nde I restriction sites , cloned from the fresh leaves of summer pokeweed PAP- RT-PCR method II genes, and T vector , were transformed into E. coli JM109 , the identification of positive subclones after Nde Ⅰ and Hind Ⅲ digestion , send Sangon sequencing . Determine positive subclones containing PAP-Ⅱ gene with restriction enzyme Nde Ⅰ and Hind Ⅲ double digestion of the positive subclones product of the target gene fragment cut plastic
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