|Title||Establishment of a complete alien tomato chromosome addition series in potato through the use of GISH and RFLP analyses|
|Author(s)||Haider Ali, S.N.|
|Source||Wageningen University. Promotor(en): E. Jacobsen; R.G.F. Visser; M.S. Ramanna. - S.l. : S.n. - ISBN 9789058083869 - 96|
|Publication type||Dissertation, internally prepared|
|Keyword(s)||aardappelen - solanum tuberosum - tomaten - solanum lycopersicum - protoplastenfusie - transgene planten - somatische hybridisatie - restrictiefragmentlengtepolymorfisme - geslachtskruising - potatoes - solanum tuberosum - tomatoes - solanum lycopersicum - protoplast fusion - transgenic plants - somatic hybridization - restriction fragment length polymorphism - intergeneric hybridization|
|Categories||Plant Breeding and Genetics (General)|
Producing somatic fusion hybrids between distantly related non-crossable taxa opened the possibilities for the introgression of useful genes or chromosomes from various species to cultivated crops. The molecular biological techniques like genome analysis by molecular markers, amplified fragment length polymorphism (AFLP) restriction fragment length polymorphism (RFLP) and in situ hybridization (ISH) have enabled the identification of genomes and individual chromosomes with great accuracy and efficiency. These techniques have surpassed all the difficulties that were faced in conventional breeding or classical cytogenetics in identification of desirable chromosomes and genes.
The main aims of the present series of investigation, were the following; a) to complete the series of monosomic alien tomato addition lines by identifying the five chromosomes (3,5,7,9,11) that were not identified in a previous investigation; b) to gain more insight into the possible phenotype in a nullisomic potato background of a known genetic locus (amylose-free) present on one of the monosomic addition chromosome; c) to explore the possibilities of increasing the prospects of intergenomic recombination between potato and tomato chromosomes by crossing with a bridge species and d) to test whether homoeologous recombination segments can be cytologically detected, i.e., through GISH, so that the alien segments can be monitored in individual potato chromosomes for the purpose of introgression of tomato encoded traits in potato.
For this purpose, basically a combination of RFLP and GISH analyses was used to identify and prove the presence of an alien tomato chromosome in potato. Where all the monosomic addition lines were identified, an additional disomic line was also identified for chromosome 11 by GISH analysis. Meiotic analysis of this disomic addition had a tendency for precocious separation of the paired homologous alien chromosomes at metaphase I and anaphase I stages. There is no definite answer for such a behaviour but a higher ploidy level of potato is reasoned as one cause. This high ploidy level is also a drawback for the phenotypic expression of alien tomato genes. To study phenotypic expression, an attempt was made to introduce the amylose-free wildtype locus of tomato ( Amft ), which is involved in the synthesis of amylose during starch formation, into a nulliplex ( amfp4 ) genotype of potato. The Amft is present on chromosome 8 of tomato that is homoeologous to chromosome 8 of potato. The idea was to complement tomato chromosome 8 for potato chromosome 8 and see its effect on starch formation and synthesis. Unfortunately, the complementation for chromosome 8 of potato was not achieved through chromosome 8 ( Amft ) of tomato because of its too low transmission frequency. Despite this absence, two types of starch phenotypes were attributed to the simplex Amfpamfp3condition, clearly indicating variable expression of the wildtype Amfp -allele in different genetic backgrounds. The two starch phenotypes observed consisted of blue or red starch granules with a blue core. However, previous investigations by our group have shown that very little, if any, homoeologous recombination occurs between the chromosomes of potato and tomato. In order to overcome the lack of recombination between potato and tomato genomes, the use of a "bridge species", Lycopersicon pennellii was tested. For this purpose, the tetraploid potato (+) tomato fusion hybrid was successfully crossed with the diploid L. pennellii which gave rise to a trigenomic hybrid. The trigenomic hybrid consisting of one genome each of potato, tomato and L. pennellii indeed showed trivalent formation at metaphase I stage during microsporogenesis but it was highly sterile. After chromosome doubling through tissue culture of this triploid a typical hexaploid was produced. Despite the expected normal meiotic chromosome pairing, the hexaploid also proved to be sterile.
In order to test whether it is possible at all to cytologically identify recombinant segments, an investigation was carried out by using F 1 hybrids between L. esculentum and L. pennellii and its backcross progeny. Because these two species are relatively closely related the homoeologous chromosomes paired and crossing over as well as recombinant segments were certainly expected in the meiotic product of their F 1 hybrids. A detailed GISH analysis of both mitotic and meiotic chromosomes indicated that in the case of small chromosomes such as found in tomato and L. pennellii it is not possible to differentiate recombinant segments in a comparable way to those found in hybrids and their backcrosses in species with large chromosomes (e.g., Alstroemeria and Lily). However, in case of tomato x L. pennellii hybrids it was possible to visualize and differentiate the heterochromatic region around the centromeres of the chromosomes and we could differentiate between the chromosomes to these two genomes following the GISH hybridization pattern. In the absence of cytological detection methods to prove introgression, an alternative has to be used such as RFLP and AFLP molecular mapping methods.
It might be concluded from the series this investigations that a) the fusion hybrids could be used for backcrossing; b) potato genotypes do retain all alien tomato chromosomes to a reasonable extent (i.e., no preferential elimination) and that these alien chromosomes are transmitted to progenies; c) there is very little or no homoeologous recombination between potato and tomato chromosomes; d) the use of the bridge species, L. pennellii can facilitate exchange of genetic material between potato and tomato after sterility problems have been solved. In principle, potato (+) tomato fusion hybrids can be used for introgression, though the process to come to it is laborious.