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molecular Genetics

molecular genetics (GB)

Term for a sub-area of molecular biology to research the structure of the genome (the entirety of the genetic material in an organism) and the function and interaction of genes (which is in the chromosomes of each cell nucleus on the DNA strand localized hereditary factors). Genetic engineering or Genetic Engineering is the application of genetic methods in practice and mostly means artificial genetic manipulation with the aim of introducing new, previously unavailable properties such as resistance against fungal diseases into an existing organism through the technically feasible implantation of individual reproduced foreign genes from foreign organisms into the genome of a variety or race.

Molecular genetics - DNA strands

Polymerase chain reaction

Since the invention of the PCR method (Polymerase Chain Reaction) for duplication of the smallest DNA amounts In the 1990s, grape varieties can now be genotypically clearly defined, defined and recognized with the help of molecular genetic characterization methods. With that, the genotype in the ampelography introduced as a complement to the traditional, visual on the phenotype based ampelography can help clarify many outstanding questions. Each independent grape variety sprouted from one seed is represented by a distinctive genotype, which is characterized by the unique combination of maternal and paternal genes during fertilization and is present in the nucleus of every plant cell. Any from sexual reproduction vine (like any other individual resulting from sexual reproduction) has an individual genetic pattern that distinguishes it from all other grape varieties.

Microsatellite analysis

The so-called genetic fingerprint, which can be created with a microsatellite analysis, is used for genotypic comparisons or the identification of grape varieties and the determination of the pair of parents. The microsatellites are defined by molecular markers and can be found within the long ones DNA chain that have a different number of repeating base sequences. Given the double set of chromosomes of the vine (2n = 38), these sequences, which are repeated twice, can be of different lengths both between two homologous chromosomes and from grape variety to grape variety. These sequences can be copied and reproduced as individual fragments, so that one obtains a certain characteristic through the length of these genome fragments.

Many such fragments of several gene locations, which are always present in pairs, result in a variety-typical and characteristic variety pattern that can be compared with the fragment patterns of other grape varieties. This fragment pattern is, so to speak, a genetic fingerprint . The probability that two grape varieties have the same pattern is statistically around 1: 6 million, with only around 14,000 existing grape varieties and breeding lines a negligible probability. There is an agreement within the European vine growing institutes that it is therefore sufficient for the genotypic characterization of vine varieties to determine the genotype at six defined gene locations (microsatellites) in order to clearly define the profile of a variety.

Before the method of microsatellite analysis, comparisons and delimitations of varieties were only comparative morphology (outer shape) of the vine possible. They described the characteristics typical of the variety and tried to filter out characteristics that made it possible to identify and differentiate the varieties. These morphological catalogs of characteristics describe the phenotype the grape variety. Both approaches have advantages and disadvantages, but they complement each other wonderfully. Genetic analyzes in the laboratory only require a small amount of genetic material as the starting material, the examinations are not dependent on external conditions such as the season, plant age, health status, virus status or growing conditions at the site, which can disfigure the phenotype (the exterior) beyond recognition.

color mutations

Laboratory examinations cannot simply be carried out blindly on thousands of vines in the field, as can be done quickly and without additional costs with the trained eye of the ampelograph. Also varieties can be identified as mutant clones or somatic chimeras vegetative propagation have emerged, can still be better differentiated phenotypically. Because the color mutants of the Pinot Noir, namely Pinot gris and Pinot Blanc, strictly speaking, are not their own, but different berry skin color variations of the same grape variety. It is a specific one mutation process which leads to the loss of dark color. Somatic chimeras like Pinot Meunier (Black Riesling) or Garnacha Peluda are much easier and faster to visualize thanks to the strong leaf hair from the original varieties Pinot Noir or Garnacha Tinta differentiate than through complex genetic analysis.

Pinot Blanc, Pinot Gris, Pinot Noir

Experience has shown that these six fixed markers are sufficient for characterization by means of microsatellite analysis. For the clarification of relationships or even the delimitation of Clone and somatic chimeric you have to use significantly more markers within a variety. But there is the rare exception that two externally different ones phenotypes (Varieties) have the same genotypic profile at six markers. Something like this can occur when coming from a grape variety mutations has branched off a somatic chimera in special cell layers. Even varieties that have arisen through self-crossing or back-crossing (mother with child) can still be very similar. Especially with sibling varieties with the same parents, this can sometimes lead to failure in the purely visual attempt at identification.

Ancestry / Parenthood

Since the mid-1990s, the genetic characterization of grape varieties has been dealt with. The fingerprints of the several thousand varieties of the world's largest range of grapes in the Domaine de Vassal were used as a basis for comparison purposes Montpellier analyzed. A central European grape variety database is emerging (see also VIVC ). In the past few years, the lineage of hundreds of grape varieties has been determined or, in many cases, corrected for the information that breeders often misrepresent. Significant ampelographers regarding DNA analysis are Dr. Jean-Michel Boursiquot, Thierry Lacombe (France), Javier Ibáñez (Spain), Dr. Erika Maul (Germany), Dr. Carole P. Meredith (USA), Dr. Ferdinand Regner (Austria), Anna Schneider (Italy) and Dr. Joseph. Vouillamoz (Switzerland).

About the DNA analysis the chloroplast can also crossing direction be checked (mother and father), because these special organ cells are only from the mother places passed. In 1997 Meredith and Bowers succeeded at the University of California for the first time to determine the parenthood of a grape variety: Cabernet Sauvignon is over Cabernet Franc (Mother) x Sauvignon Blanc (Father) emerged. As a result, some important Central European leading varieties how Gouais Blanc (White Heunisch), Pinot and Savagnin ( Traminer ) be identified. The old age and the widespread distribution of these varieties have favored the spontaneous crossing and thus the emergence of many other grape varieties. Gouais Blanc / Heunisch, which was cultivated throughout Central Europe in the Middle Ages, has crossed into 80 grape varieties alone.

Crossing - Cabernet Franc (red) x Sauvignon Blanc (white) = Cabernet Sauvignon

As with the paternity test, the genetic fingerprint can now be used for the first time to reconstruct ancestry, determine relationships or crossbreeds of hybrids detect. However, a prerequisite is an extensive genetic analysis at at least 15 to 50 gene locations (markers). The Swiss biologist Dr. José Vouillamoz thinks it should be at least 30 to 60 (when identifying the Cabernet Sauvignon it was 35). Only when the potential parents are known, for example, from the breeder, are less enough. Since each child has inherited one chromosome from the father and one from the mother, each grape variety at a specific gene location must also have a characteristic with the father places and the other with the mother places have in common. This rule applies strictly, which means that failure to meet the parentage criterion usually means the end of the parentage hypothesis. The more gene locations that are included in the study, the safer it is. However, 100% proof cannot be provided, since the hypothesis can only be tested positively at a few selected, but never at all, gene locations.

If one of the parents cannot be identified because its DNA values have not yet been recorded and are not known, or because the variety may already be extinct), it is still possible to demonstrate the relationships between parent plants and the offspring of two different varieties. The phrase "not directly related" is often used in sources. This usually means that there are none between the grape varieties considered Parent-offspring relationship, that is, there is no “mother-father-child relationship”. Direct relationships between grape varieties (ie "mother-father-child") can be clearly reconstructed using the genetic fingerprint. Even with siblings or "more distant relatives" like "aunts" or "uncles" or "cousins", the relationship from the genetic profiles is hardly obvious.

genetic engineering

In the breeding Modern grape varieties with above all certain resilience have been using manipulative ones for some time genetic engineering, In this regard, the international project IGGP (International Grape Genome Program) launched. In tissue culture suspensions, foreign resistance genes with desired properties can be introduced into the genome of plant cells of certain conventional varieties. For example resistance against fungal diseases and virus or against animal pests like the dreaded phylloxera, Vine plants can be regenerated from the genetically manipulated cells, which this time are not a new variety, but have essentially retained the genetic properties of the selected variety, but supplemented by the "improving" infiltrated genes.

In practice, however, the hoped-for effects mostly failed to materialize or were unsatisfactory. In the case of woody plants, it is obviously not enough to simply insert individual genes somewhere into the genome and hope that complex and environmentally dependent properties such as fungal resistance may already be expressed. To what extent this technology will lead to success and practical application in terms of effort and costs for the vine is currently not foreseeable and is not in demand. In any case, the introduction of genetically manipulated grape variety clones would further impoverish varieties and further reduce the clone diversity to a few Clones to lead. Whether these artificially manipulated vine plants, whose genetic balance is disturbed, are in abundance vegetative propagation To prove stable in the long run cannot be answered seriously today.

wine adulteration

DNA analyzes in wine are used today with great success wine adulteration (Panties) to track down. Researchers from the institute INRA In 2002, Montpellier developed a method for DNA isolation from wine and must that can be used to differentiate pure quality wine from cheaper blends. The methodology has advanced to such an extent that unfiltered wine can be determined using the fingerprint typical of the variety whether a wine from the specified grape variety has been pressed or whether must from other grape varieties has been added. A quantitative determination of the proportions of the different types in wine is not (yet) possible, but one can reconstruct which types are present in the wine.

So far, the INRA team has identified the genetic profiles of 600 grape wines. The method does not work with bottled wines, however, because a large part of the DNA (DNA) is filtered out, and the residual concentration of remaining DNA in the finished wine is too low for genetic analysis. However, the research continues intensively. Using the method PNA FISH can be microbiologically based wine faults such as horse sweat be detected. See also on this topic under the keywords Nuclear Magnetic Resonance and Certification of vines,

Images: Ursula Brühl, Doris Schneider, Julius Kühn Institute (JKI)

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