Periodic Reporting for period 4 - CRISBREED (Multidimensional CRISPR/Cas mediated engineering of plant breeding)
Período documentado: 2022-04-01 hasta 2022-09-30
The basic principle of classical plant breeding is to combine advantageous traits in crops like disease resistance and high yield and at the same time eliminate unfavorable traits like bitter taste. For this purpose, the breeder is crossing an elite cultivar of a crop with close relatives. The genetic information coding for all traits is coded in genes, which are linearly packed like beads on a string on a smaller number of individual chromosomes. By crossing, an exchange of traits between parental chromosomes can be achieved. However, in many cases individual traits cannot be combined due to their specific position on a chromosome, especially if they are close to each other. Thus, it is often not possible for the breeders to obtain crops resistant to certain pests and the farmer has to use pesticides to protect the harvest. Also due to global warming, the heat and salt resistance of various crop plants needs to be improved, to avoid deleterious effects in future. These very important challenges for our global agriculture could be achievable by the development of specific technologies that allow controlled changes of plant chromosomes. Thus, it should be possible to change the structure of individual chromosomes as well as to the exchange segments between different chromosomes. Till now, the use of molecular scissors like CRISPR/Cas was restricted to the change of an individual and or a few genes. With this project we were worldwide the first to establish novel CRISPR/Cas based technologies for plant chromosome engineering. We were not only able to demonstrate that Mbp-sized intrachromosomal inversions can be obtained but that also arms between different chromosomes can be exchanged. Moreover, we also developed a new technology for the controlled elimination of specific tissues, “CRISPR-Kill”, which could be used e.g. to eliminate specifically cells that produce toxins. Thus, we established a novel kind of breeding that will help us to cope with the challenges of the 21th century. Closely linked traits can now be separated and genetic exchange can be induced in regions of the plant genome that were inaccessible for breeding before.
The main outcome of this project is that as new technology CRISPR/Cas mediated chromosome engineering was set up for plants. All in all, three major breakthroughs could be achieved: Chromosomal inversions, chromosomal translocations and tissue engineering by genome elimination.
Chromosome engineering is of special interest for plants as basic problems of crop breeding can be solved this way. Till now it was not possible to separate traits if the respective genes are located in close proximity on the same chromosome. Moreover, inversions hinder the exchange of genetic information in specific regions of chromosomes. For both problems we were able to develop proof of concept solutions: By exchanging chromosome arms we are able to separate genes that were genetically linked before. On the other side it should also be possible to bring genes in close proximity that should be transferred together to the next generation. By long read sequencing of crop genomes it only became clear in the last five years that a large number of genome variations between crop varieties is due to large inversions. In the inverted regions no genetic exchanges are possible, which is a big drawback for breeding. We demonstrate that by inducing controlled inversions we can not only bring back genetic change to these regions. Moreover, we were able to block genetic exchange on almost an entire chromosome by inducing inversions. Thus, genes can be genetically linked for the breeding process, which was not possible before.
By now - based on our results in the model plant Arabidopsis – chromosome engineering is already applied by the community to solve breeding problems with crop plants and large inversions have already been reported for corn and rice.
By serendipity we realized that we are able to kill cells by induced double strand breaks in functional repeats. Using promoters specific for individual organs we were able to develop a novel way of tissue engineering. Thus, we were able to block in a controlled the formation of specific organs like petal or side roots. “CRISPR-Kill” could not only be an attractive new tool for controlling secondary metabolism in plants it might also be useful for tissue engineering in all different kinds of multicellular organisms.
Chromosome engineering is of special interest for plants as basic problems of crop breeding can be solved this way. Till now it was not possible to separate traits if the respective genes are located in close proximity on the same chromosome. Moreover, inversions hinder the exchange of genetic information in specific regions of chromosomes. For both problems we were able to develop proof of concept solutions: By exchanging chromosome arms we are able to separate genes that were genetically linked before. On the other side it should also be possible to bring genes in close proximity that should be transferred together to the next generation. By long read sequencing of crop genomes it only became clear in the last five years that a large number of genome variations between crop varieties is due to large inversions. In the inverted regions no genetic exchanges are possible, which is a big drawback for breeding. We demonstrate that by inducing controlled inversions we can not only bring back genetic change to these regions. Moreover, we were able to block genetic exchange on almost an entire chromosome by inducing inversions. Thus, genes can be genetically linked for the breeding process, which was not possible before.
By now - based on our results in the model plant Arabidopsis – chromosome engineering is already applied by the community to solve breeding problems with crop plants and large inversions have already been reported for corn and rice.
By serendipity we realized that we are able to kill cells by induced double strand breaks in functional repeats. Using promoters specific for individual organs we were able to develop a novel way of tissue engineering. Thus, we were able to block in a controlled the formation of specific organs like petal or side roots. “CRISPR-Kill” could not only be an attractive new tool for controlling secondary metabolism in plants it might also be useful for tissue engineering in all different kinds of multicellular organisms.
For the first time worldwide, we were able to establish heritable plant chromosome engineering. We also developed a novel CRISPR/Cas way of tissue engineering.