caa a22 4500 463160843 CHVBK 20180406164747.0 cr unu---uuuuu 170326e20070601xx s 000 0 eng 10.1007/s10535-007-0053-5 doi (NATIONALLICENCE)springer-10.1007/s10535-007-0053-5 Production of human papillomavirus type 16 E7 oncoprotein fused with β-glucuronidase in transgenic tomato and potato plants [Elektronische Daten] [J. Bříza, D. Pavingerová, J. Vlasák, V. Ludvíková, H. Niedermeierová] The human papillomavirus type 16 (HPV 16) oncogene E7 fused with the gene for β-glucuronidase (gus) was used in plant transformation experiments. The E7 gene modified for lower cancerogenicity and fused with the 5′ end of the gus in cassettes with cauliflower mosaic virus 35S promoter and transcription terminator produced high contents of fusion proteins in potato protoplasts. Expression vectors harbouring E7 fusion cassettes were used for Agrobacterium tumefaciens LBA4404 mediated transformation of either potato (Solanum tuberosum L. cv. Bintje) or tomato (Lycopersicon esculentum Mill. cv. Moneymaker). A fusion gene was found in all rooted regenerants using polymerase chain reaction with primers providing amplified fragments from E7 and gus genes. GUS activity was revealed in all regenerants obtained. Nevertheless, the level of GUS expression in different constructs varied much more than in transient expression experiments with potato protoplasts. Especially, expression level in plants carrying vectors with the whole E7 gene fused with gus was lowered by 2-3 orders of magnitude comparing with fusion of the first 41 codons of E7 and gus. Southern hybridisation of 18 tomato and 23 potato regenerants revealed mostly multiple tandem integration of T-DNA into the plant genome and Western blot proved the presence of the fusion protein in 9 tomato and 11 potato plants out of 41 tested individuals. Institute of Experimental Botany, ASCR, 2007 E7/GUS fusion protein nationallicence HPV 16 nationallicence Lycopersicon esculentum nationallicence Solanum tuberosum nationallicence transformation nationallicence 2,4-D : 2,4-dichlorphenoxyacetic acid nationallicence CaMV : cauliflower mosaic virus nationallicence ECL : enhanced chemoluminiscence nationallicence GUS : β-glucuronidase nationallicence HPV 16 : human papillomavirus type 16 nationallicence MES : 2-(N-morpholino)ethanesulphonic acid nationallicence MOPS : 3-(N-morpholino)propanesulfonic acid nationallicence MUG : 4-methyl-umbelliferyl-β-D-glucuronide nationallicence NPTII : neomycin phosphotransferase II nationallicence ORF : open reading frame nationallicence PBS : phosphate buffered saline nationallicence PEG : polyethylene glycol nationallicence SDS-PAGE : sodium dodecyl sulphate polyacrylamide gel electrophoresis nationallicence Bříza J. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut Pavingerová D. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut Vlasák J. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut Ludvíková V. Institute of Hematology and Blood Transfusion, U nemocnice 1, CZ-12820, Praha 2, Czech Republic aut Niedermeierová H. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut Biologia Plantarum Kluwer Academic Publishers-Consultants Bureau 51/2(2007-06-01), 268-276 0006-3134 51:2<268 2007 51 10535 https://doi.org/10.1007/s10535-007-0053-5 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10535-007-0053-5 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Bříza J. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut NATIONALLICENCE 700 1- Pavingerová D. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut NATIONALLICENCE 700 1- Vlasák J. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut NATIONALLICENCE 700 1- Ludvíková V. Institute of Hematology and Blood Transfusion, U nemocnice 1, CZ-12820, Praha 2, Czech Republic aut NATIONALLICENCE 700 1- Niedermeierová H. Institute of Plant Molecular Biology, Biological Centre of the Academy of Sciences of the Czech Republic, Branišovská 31, CZ-37005, České Budějovice, Czech Republic aut NATIONALLICENCE 773 0- Biologia Plantarum Kluwer Academic Publishers-Consultants Bureau 51/2(2007-06-01), 268-276 0006-3134 51:2<268 2007 51 10535 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer