الفهرس 
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Abstract
This study is part of the project, entitled “Genomic analysis of the relationships between the potato’s response to photoperiod, vegetative period, and late blight résistance“, two points were studied, the first point was about a gene that plays a role in pathogenicity in the pathogen (Phytophthora infestans) called, Phytophthora infestans cleavage-associated kinase 1 gene (Pick 1) while the second point was about a new gene could be play a role in resistance to late blight disease in the host (Potato) called, The pathogenesis-related protein, isoform b2 (pr1-b2), the obtained results throughout this study could be summarized as follow:
1. Detection of allelic variants of a gene of Phytophthora infestans that plays a role in pathogenicity:
Total genomic DNA were extracted from P. infestans and DNA concentration was estimated visually in agarose gels The samples were routinely diluted to 50 ng/µl by using TE buffer for PCR amplification of a portion of the Pick 1 gene. The primers Pick1-for and Pick1-rev amplified 1120–bp-fragments from genomic DNA of 18 P. infestans isolates that appeared on agarose gels as a single band thus confirming the presence of a single genetic locus. Alleles were marked by single nucleotide substitutions (SNPs), no insertions, deletions and introns were apparent or detected. A total of 9 SNPs (single nucleotide polymorphisms) were detected within the nucleotide sequence of Pick 1 fragments, these could be assigned to a total of two different alleles, Pick1a and Pick1b. Seven isolates were homozygous for Pick1a, two isolates were homozygous for Pick1b and nine isolates were heterozygous Pick1a/b. The single sequence represented by the Genbank accession, AY093611, coincided with allele Pick1a at seven and with Pick1b at two imperfectly consecutive SNP positions (tentative recombinant allele Pick1c), this allele was however not present among our isolates.
Two allele-specific MAMA primers were designed using the T/C dimorphism at position 390, therefore albeit highly allele-specific primers for PCR, this was achieved by applying primers possessing a mismatch nucleotide at position -3 from the 3’-terminus, although these primers cannot differentiate between allele Pick1a and the putative allele Pick1c, it was assumed that allele Pick1c represented by AY093611 would not occur in Austria. The development of this method of rapid SNP detection and the distinction of individual alleles at this locus was applied to the genetic fingerprinting of natural P. infestans populations in Austria during 2003 to 2005 and used to investigate the association of specific alleles with differing degrees of P. infestans zoospore formation and the development of late blight disease on host.
The isolates of P. infestans tested in the study proved to be of the A1 and A2 mating types but The A1 mating type was more frequent in 2003 and 2005 than A2 and the opposite was true in 2002 and 2004, across all geographic regions of Austria sampled This suggests that the Austrian P. infestans populations may belong to the ”new” type , which represents both clonal and sexual reproduction as can be inferred from the widespread and abundant occurrence of both the A1 and A2 mating types within our representative samples of the Austrian populations.
2. Detection of allelic diversity at the potato pr1-b2 locus and test for association with resistance to late blight:
pr1-b2 was discovered at 2006 and only one genetic sequence has been available from public databases. This sequence was used to design PCR primers that readily amplified several DNA fragments from genomic DNA was extracted from 10 different potato cultivars (diploid and tetraploids hybrids), Alleles were detected by single nucleotide polymorphisms (SNPs) and one small insertion of three nucleotides, no introns were apparent.
Several SNPs (single nucleotide polymorphisms) were detected within the nucleotide sequence of pR1-b2 gene. Work continued with the two cultivars MF-II and TPS 67 that were parents of a 200-progeny population segregating for late blight resistance. The putative alleles as assumed the patterns of SNPs were named for the purpose of the investigation as MF-II-1, MF-II-2, MF-II-3, MF-II-4, TPS67-1, TPS67-2, TPS67-3, and TPS67-4.
Six allele-specific primers were designed based on informative SNPs. The primers reliably amplified the expected fragment only when the respective allele, as detected by sequencing, was present. they could be used to test for association with late blight resistance in the MF-IIxTPS67 population.
The markers obtained with these primers were also used for genetic mapping which performed of their relative positions and for further characterization of the locus or loci in the potato genome that may correspond to the pr1-b2 gene from S. phureja.These markers mapped to different anonymous linkage groups and due to the limited size of the map for this tetraploid population it was not yet possible to infer the identity of these anonymous linkage groups with any of the 12 chromosomes of Solanum. Therefore, it can be concluded that by the method of genetic mapping no genetic locus of S. tuberosum with close similarity to pr1-b2 from S. phureja was detectable that may be strongly associated with the late blight resistance as seen in the MF-IIxTPS67 population.
Prior to the presented investigation, the late blight resistance genotypes of MF-II and TPS67, the parents of the population, had been inferred from the resistance of each parent to different isolates of P. infestans and from the pattern of segregation of the resistance to the same isolates in the MF-IIxTPS67 progeny of 200 clonal siblings. It was possible to phenotype all progenies and parents of the population with isolates that could not break down any of the R genes and with isolates that broke down either one or both of the R genes.
These progenies for which the resistance genotype were partly used for genotyping with the PCR markers developed on pr1-b2 as outlined above
The two sets of data, resistance genotype and marker genotype, were used to search for potential associations of these molecular markers with any of the resistance genotypes. No PCR marker was entirely associated with one resistance class, i.e., there was no marker that was exclusively present in all plants of one specific resistance class and there was no marker that was entirely missing in one class. At the same time, the figures of markers present and absent obtained for every marker within every resistance class showed the same distribution as was expected for each marker from its individual genetic status. Therefore, it is safe to conclude that none of the PCR markers tested were strongly associated with any of the resistance genotypes inferred within the MF-IIx TPS67 population.
In conclusion, it can be deduced that the pr1-b2 locus does not play any important role for race-specific resistance determined by the major resistance loci R1(MF-II) and RT. It could, however, still play a role in additively controlled race-nonspecific resistance. Additional experiments and calculations will be necessary to test this alternative hypothesis.