Biovigilance: Conceptual Ideas for the use of COIS(Copyright Owner Identification System) in GMO-Tracing

The GMO-COIS is a new technology which was developed by ATG:biosynthetics to identify GMOs by using artificial nucleic acid sequences. These sequences are employed for the reliable and explicit labelling and thus identification of GMOs. For this purpose, artificial sequences are combined with genes, which are to be introduced into a recipient organism. By using primers that bind to specifically designed sequences the constructs can be amplified by Polymerase Chain Reaction. The resulting (labelled) amplification products are transferred to microarray-chips onto which probes with known sequence identity have been immobilized. Binding of an artificial sequence to its complementary sequence on the chip allows the identification of the artificial sequences. This also requires a database containing all assigned sequences, e.g. represented by a sort of a bar code. There is a multitude of applications for this technique:

• A new variety can be branded with artificial sequences. Stable integration of the sequences into the genome of the progeny makes it possible to identify a GMO generations later. On one hand, this system can be implemented to guarantee or confirm a breeder’s property right. This is even the case for vegetatively propagated plants. On the other hand, consumer protection can be improved.
• Control of cells, which are inserted for clinical trials, e.g in gene or cell therapy studies.
• Control of micro-organisms in fermenters.
• Detection of horizontal gene transfer from GMOs to their wild-type counterparts in the field.
• Unequivocal identification of plants with traits and genes that should not enter the food chain such as plants used for the production of biopharmaceuticals (Molecular Pharming).

Document Downloads related to the talk at the Conference on Plant-made Pharmaceuticals in Quebec City - March 16-19, 2003 Part1 Part2 Part3 Part4

Initiative for introducing a highly efficient GMO tracing system

Today I would like to present an alternative approach to the monitoring of transgenic organisms and transgenes in general. The alleged perils and potential benefits that stem from transgenes and/or transgenic organisms are in the news frequently. Uncontrolled spread of transgenes to the food chain is a major concern to the public, but also to a number of scientists and many governments (see EU legislation, directive 2001/18/EC). What is asked for in debates and scientific symposia (e.g. the 7th International Symposium on the Biosafety of Genetically Modified Organisms - www.worldbiosafety.net) is an effective, reliable, and, ideally, universal system for detecting, monitoring, and tracking transgenes in their daily use. Standard procedure is to detect transgenes or their protein products individually by analysis in high-throughput set-ups, e.g. based on PCR technologies, ELISA, etc. To date only a few (biochip-based) HTS systems with the potential of simultaneously detecting multiple transgenes exist. One shortcoming of various tests is that they assess the presence of markers in the carrier vector (promoter or terminator sequences, antibiotic resistance) rather than identifying the concrete nature of the transgene itself. In addition, different tests are offered that make cross-correlation of results difficult. What would help overcome these constraints is a reliable generalized and standardized approach to tagging transgenes (in organisms) a priori.

As a next step, ATG would provide the arrays/biochips needed to detect tagged DNA from GMOs. There are at least two options here: for in-house use you can purchase a biochip spotted with a selection of oligonucleotides necessary to profile your transgenes. For a more general use biochips with a higher oligonucleotide density are available to detect a host of transgenes, e.g. for monitoring investigations carried out by government bodies.

Join the initiative and take full advantage of all its benefits! We look forward to your feed-back on this initiative! [info]

To start with, you will obtain a set of tags in the shape of a bacterial colony (stab culture) containing a unique mixture of degenerate artificial DNA sequences. Either the customer himself isolates individual clones for use as unique tags by standard molecular biology techniques. Alternatively, you can commission us to do all the screening, sequencing, and associated lab work. You would then receive a collection of quality-checked and documented clones each carrying a different tag. GMO-COIS uses completely artificial sequences for its tags. These sequences are unique and have little or no similarity to any sequence encountered in nature. Bacterial colonies carrying degenerate DNA sequences, i.e. the actual tags, are available to different customer groups at the following rates:

Prices on request!

How does the system work? An infocode-id is constructed by designing a DNA cassette consisting of:

• a random, completely artificial "bar code-like" sequence. This sequence corresponds to a database entry with detailed information, e.g. on (a) the nature of the transgene: type of DNA construct, organism used for transfer, etc. (b) name of person or business authoring the GMO, (c) copyright information such as date and place of issue, etc., (d) variants of the transgene or the genetically tagged organism, (e) documented references for the transgenic organisms (links to web sites, data bases, publications, accession numbers, etc.). The random assignment, or in other words lack of systematic correlation, of a sequence to a database file makes the tagging procedure very resistant to forgery.
• flanking sequences at the 5’- and 3’-end are used to amplify the bar code cassette. General primers amplify information accessible to authorities or other authorized parties for reasons of monitoring, etc. Specific primers known only to the copyright holder allow identification of intellectual property.
• The artificial DNA sequence represents information that can be encrypted so that only with the appropriate key access to the clear text is possible. This makes sure that sensitive information is only passed on to authorized persons.
• The system also allows amplification of all information between sets of flanking primers. This way, complete transgene constructs can be amplified by traditional PCR. Hybridization of PCR product to probes on an appropriate biochip generates a unique profile for the construct.

Imagine having a system that works somewhat like the bar codes in supermarkets where an item and the information pertaining to it are "encoded" in a string of parallel lines. ATG:biosynthetics has set out to introduce such a system by the name of GMO-COIS (Genetically modified organism copyright-identification-system). Here, patterns of artificial DNA serve as bar code-like tags to clearly identify any transgene. GMO-COIS has the potential to become a gold standard for transgene tagging and monitoring if accepted as a binding standard. It offers the detection of up to millions of tag sequences integrated on high-density arrays (biochips). Data on tagged transgenes would then be stored and catalogued in a central data base, ideally on the basis of a voluntary contribution or a mandatory registration enforced by the responsible government bodies. Essential information on a transgene, e.g. primer sequences for amplification, should be made available to federal research laboratories and government organizations. Access to sensitive data could be limited to authorized parties. Data flow and exchange between interested parties becomes a lot easier by transferring access rights to business partners. A state- or government-appointed controlling authority or, alternatively, a service provider could be entrusted with updating and maintaining the data base.

The advantages of such a system are:

• (Cost-efficient) central registration and administration of author and GMO data
• Access to most recent data on transgenic organisms
• Automated detection and array synthesis can be implemented: Possibility of feeding new data to remote array synthesizers where updated arrays can be created
• Ultra-dense integration of different author data onto a microarray system (millions of authors and data)
• Generalized detection method ("one tube") in contrast to individualized detection of GMOs (numerous different analyses).
• Minimum need for and consumption of sample material (reaction volumes in the microliter range)

Applications:

• Quality assurance: Assess the genetic integrity of your transgenic organisms (stab cultures, pro- and eucaryotic strains, etc.). Determine contamination with and nature of undesired transgenes.
• In-house records: Keep track of all transgenes created or currently used in your research, brand new varieties with artificial sequences. Due to the stablle integration of the sequences into the host genome progeny of a GMO can be identified many generations later.
• Copyright identification: protect your business against unauthorized use and marketing of intellectual property. Breeders have a means to verify their property rights.
• Tracking of GMOs in the food and agro industry form source to customer: Unequivocally determine the origin of a transgenic organism (producer), its distribution routes, etc. The food and agro industry could take advantage of the system that allows complete tracking of distribution routes of transgenic bacteria, plants and animals all the way from their breeding/creation and release into the environment all the way to their final destination, either in laboratories or with the consumer. This will help improve consumer protection.
• Control: unequivocal identification of transgenes that should not enter the food chain, e.g. in plants used for molecular pharming of biopharmaceuticals.

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