General informationNotification NumberB/DE/04/160Member State to which the notification was sentGermanyDate of acknowledgement from the Member State Competent Authority13/10/2004Title of the ProjectEvaluation of transgenic potato as bioreactor for production of recombinant spider silk under field conditionsProposed period of release:01/04/2005 to 31/10/2005Name of the Institute(s) or Company(ies)Institut für Pflanzengenetik und Kulturpflanzenforschung Gatersleben (IPK), Gatersleben, Corrensstr. 3, D-06466 Gatersleben;
3. Is the same GMPt release planned elsewhere in the Community?NoHas the same GMPt been notified elsewhere by the same notifier?NoGenetically modified plantComplete name of the recipient or parental plant(s)
2. Description of the traits and characteristics which have been introduced or modified, including marker genes and previous modifications:For the deliberate release potato plants (Solanum tuberosum), variety Albatros, were modified by genetic engineering. The plants were transformed by Agrobacterium-mediated gene transfer. The transferred genes of interest are coding for spider silk proteins and elastin-like proteins (MaSpI-100ELP, MaSpII-100ELP and SO1-100xELP). Transgenic potato plants as bioreactors for the production of spider silk proteins under field conditions will be evaluated.
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MaSpI and MaSpII are spider silk proteins from Nephila clavipes. SO1 is a synthetic spider silk protein and mimics the repetitive part of MaSpI from Nephila clavipes. 100xELP is a synthetic gene and is homologous to human elastin. MaSpI-ELP, MaSpII-ELP and SO1-ELP are fused at the N-terminus to the Vicia faba legumin A signal peptide sequence. This causes the translation at the ER associated ribosomes and the transport into the Endoplasmic Reticulum. Between the sequences coding for MaSpI, MaSpII and SO1, respectively, and the sequences coding for 100xELP a sequence coding for the human c-myc-tag was inserted. The c-myc-tag is a short amino acid sequence out of the human C-MYC protein. It is used for the immunochemical detection of the fusion proteins and has no enzymatic activity. The sequences coding for the fusion proteins are C-terminally fused to the sequence coding for the ER retention signal KDEL. This causes retention of the recombinant proteins in the ER. The expression of the fusion proteins is regulated by the CaMV35S promoter, an ubiquitous promoter. This promoter is regulates the expression of 35S transcripts of Cauliflower Mosaic Virus. For termination of transcription the CaMV35S untranslated region at the 3énd is used.
To differentiate between transformed and untransformed cells a gene coding for kanamycin resistance was introduced into the genome of the transgenic potato plants. The resistance gene is regulated by the nopaline synthase promoter and terminator from Agrobacterium tumefaciens. The Kanamycin resistance gene codes for a well characterized enzyme, the neomycin phosphotransferase. There is no hint for toxicity. Several transgenic plants containing the neophosphotransferase coding gene were evaluated by the US Food and Drug Adsministration (FDA) and by the US Environmental Protection Agency and characterized as completely save. Those plants are generally integrated into the US agriculture. Some lines contain the nptIII gene, that could be expressed only in bacteria, but not in plants. The nptIII gene causes resistance against Kanamycin, Neomycin and Amikacin. Amikacin is an important antibiotic of value in therapy. Taking into account, that (i) the probability of horizontal gene transfer from plants to bacteria is very low and (ii) there is no selection pressure at the area of deliberate release it could be assumed, that the frequence of nptIII caused resistance in microorganisms will not be assumed by the deliberate release.Genetic modification3. Type of genetic modification:Insertion; In case of insertion of genetic material, give the source and intended function of each constituent fragment of the region to be inserted:Description of composition and origin of the transgene: -CaMV35S promoter from Cauliflower Mosaic Virus: promoter for expression of the gene of interest; LeB4-signal peptide from Vicia faba L.: signal peptide for transport of transgenic protein of interest into the Endoplasmatic Reticulum; spider silk gene MaSpI from Nephila clavipes: structure-protein of spider silk; spider silk gene MaSpII from Nephila clavipes: structure-protein of spider silk; synthetic spider silk gene SO1 with homology to MaSpI from Nephila clavipes: structure-protein of spider silk; c-myc-tag from Homo sapiens: Immunochemical detection with antibodies in western blot, without enzymatic activity and without DNA-adsorbtion activity; synthetic elastin 100xELP with homology to human elastin: structure-protein of connective tissue; ER-Retionssignal KDEL from Homo sapiens: retention of heavy chain binding protein out of Endoplasmic Reticulum; polyadenylation signal from Cauliflower Mosaic Virus: termination of transcription of gene of interest.
Description of composition and origin of the T-DNA: Nos promoter from Agrobacterium tumefaciens: promoter for expression of nptII gene; nptII-gene from E. coli: gene coding for the Neomycin phosphotransferase; part of Ocd-gene from Agrobacterium tumefaciens: non- functional part of the gene coding for the Ornithin cyclodeaminase; Nos terminator: termination of transcription; part of geneIII from phage M13: non-functional part of the gene; part of lacZ-gene from E. coli: non-functional part of the gene
The following parts outside of the T-DNA are also transferred: OriV from E. coli: not functional in plants; nptII-gene from Streptococcus faecalis: not functional in plants; trfA-gene for replication insite of E. coli and Agrobacterium tumefaciens: not functional in plants.6. Brief description of the method used for the genetic modification:The genetic modification was brought about by Agrobacterium-mediated gene transfer.7. If the recipient or parental plant is a forest tree species, describe ways and extent of dissemination and specific factors affecting dissemination:not applicableExperimental Release1. Purpose of the release:The purpose of the release is to evaluate potatos as bioreactors for the production of spider silk proteins.2. Geographical location of the site:The field release will be at IPK Gatersleben, Saxony-Anhalt3. Size of the site (m2):20004. Relevant data regarding previous releases carried out with the same GM-plant, if any, specifically related to the potential environmental and human health impacts from the release:No data regarding previous releases related to potential environmental and human health impacts.Environmental Impact and Risk ManagementSummary of the potential environmental impact from the release of the GMPts:Due to the absence of sexual compatible wild species and in the light of the modification carried out, no obvious environmental risks can be seen.Brief description of any measures taken for the management of risks:The release site will be monitored during and after the release. Potato lants not intended to be used for experimental purposes will not be grown in the vivinity of the release site.Summary of foreseen field trial studies focused to gain new data on environmental and human health impact from the release:not applicableFinal reportEuropean Commission administrative informationConsent given by the Member State Competent Authority:Yes10/05/2005 00:00:00Remarks: