Genetic fingerprinting of Hungarian sour cherry cultivars

Transkriptio

1 Júlia Halász, Attila Hegedűs Genetic fingerprinting of Hungarian sour cherry cultivars OTKA PD78124 János Bolyai Scholarship, HAS Department of Genetics and Plant Breeding, Corvinus University of Budapest, Hungary Institute for Extension and Development, University of Debrecen, Hungary

2 Future tasks Memories from San Michele all Adige (Trento), 1th OCTOBER 212 Sour cherry selection is an ongoing task in North-Eastern Hungary Pipacs 1 : identification of mislabelled genotypes by using molecular markers (gene-specific and SSR) Developing molecular markers for dominating specific compounds in sour cherry selections Animal and clinical studies to characterize the efficiency of different sour cherries in preventing and treating of several diseases Self-(in)compatibility studies in sweet and sour cherries, crop evolution

3 The aims of the study Genetic fingerprinting of Hungarian sour cherry germplasm Working out SSR based discrimination for phenotypically similar accessions Checking putative clonal status of some selections Analyse the relationships of morello and amarelle-type sour cherries Finding new markers as well as new ways of marker application in sour cherry

4 Montmorency Csengődi Pándy Cigánymeggy Pipacs 1 Fanal

5 From yellow to the black

6 Plant material Accession Pedigree Place of origin Self-(in)compatibility Cigánymeggy 44 Carpathian Basin SC Cigánymeggy 7 Carpathian Basin SC Csengődi Akasztó SC Debreceni bőtermő Debrecen SC Du-1 Dunavecse SI Érdi bőtermő Pándy Nagy angol Érd SC Érdi jubileum Pándy Eugenia Érd SC Fanal Schattenmorelle Unknown Germany SC Favorit Pándy Montreuilli Érd SC Kántorjánosi Mátészalka SC Korai pipacs Pándy Császár Érd SC Maliga emléke Pándy Eugenia Érd SC Meteor korai Pándy Nagy angol meggy Érd SC Montmorency France SC L Clone or hybrid of Érdi b. Újfehértó SC N2 Clone or hybrid of Érdi b. Újfehértó SC N212 Clone or hybrid of Érdi b. Újfehértó SC N3 Clone or hybrid of Érdi b. Újfehértó SC N4 Clone or hybrid of Érdi b. Újfehértó SC N5 Clone or hybrid of Érdi b. Újfehértó SC Pándy 279 Carpathian Basin SI Pipacs 1 Kecel SC Piramis M221 Meteor korai Érd??? Újfehértói fürtös Újfehértó SC

7 Methodology 1 SSR primer pairs CPSCT 21 Mnejja et al., 24 CPSCT12 Mnejja et al., 24 BPPCT 38 Dirlewanger et al., 22 BPPCT 2 Dirlewanger et al., 22 BPPCT 4 Dirlewanger et al., 22 3 primer pairs for S-genotyping PaConsI-F and R Sonneveld et al., 23 PaConsII-F and R Sonneveld et al., 23 EM-PC2consFD and EMPC3consRD Sutherland et al.,24 BPPCT 37 Dirlewanger et al., 22 BPPCT 39 Dirlewanger et al., 22 BPPCT 4 Dirlewanger et al., 22 ASSR17 Xu et al., 24 ASSR63 Xu et al., 24 ABI PRISM 31 Genetic Analyzer, Genotyper 3.7 software and the GS5 LIZ size standard (Applied Biosystems) In silico DNA sequence analysis

8 SSR fingerprints of Korai pipacs and Pipacs 1 julia.halasz@uni-corvinus.hu

9 SSR fingerprints of Amarelle-type cherries

10 Similarity Pipacs 1 Újfehértói fürtös Debreceni bőtermő Kántorjánosi Dendrogram of Hungarian sour cherries Pándy 279 N2 N3 N4 N5 N L Érdi bőtermő 5 51 Meteor korai Piramis Korai pipacs Maliga emléke Favorit Érdi jubileum Du-1 Fanal 81 Montmorency Cigánymeggy Cigánymeggy C. 44 Csengődi julia.halasz@uni-corvinus.hu

11 Preliminary results of S-genotyping 15 bp 1 bp 75 bp 5 bp 25 bp S 36b S 1 S 6 S 4 S 36b2 S 9 S 1 F K Éj Mk Me Du C7 Cs Úf S S 4 S 4 S 4 S 4 S 4 S 4 S 4 4 S 36a S 36b2 S 36a S26 S 1 S 6m S 6m S 6m S 6m S 6m S 6m S 6m S S 6 9 Pá C4 Db Pi N2 N3 N4 N5 N21 L Éb P1 Kp Fan M

12 Identification of S 36b2 in Pipacs P. cer. Pipacs 1 ATGACCAAGTAACTATTCAAACCCAAAGAGGCCCAGTAATTGCAGGGGGTCGCTATTTGATTCAAGGAAAGTGGTATGTATTGATTTTTTTCTCACTTACTCTTTAGCATTTAGTTTAAA 12 P. cer. S-RNase-S36b2 ATGACCAAGTAACTATTCAAACCCAAAGAGGCCCAGTAATTGCAGGGGGTCGCTATTTGATTCAAGGAAAGTGGTATGTATTGATTTTTTTCTCACTTACTCTTTAGCATTTAGTTTAAA 12 P. cer. S36b3-RNase ATGGCCAAGTAACTATTCAAACCCAAAGAGGCCCAGTAATTGCAGGGGGTCGCTATTTGATTCAAGGAAAGTGGTATGTATTGATTTTTTTCTCACTTACTCTTTAGCATTTAGTTTAAA 12 P. cer. S36b-RNase ATGGCCAAGTAACTATTCAAACCCAAAGAGGCCCAGTAATTGCAGGGGGTCGCTATTTGATTCAAGGAAAGTGGTATGTATTGATTTTTTTCTCACTTACTCTTTAGCATTTAGTTTAAA 12 P. cer. S36a-RNase ATGGCCAAGTAACTATTCAAACCCAAAGAGGCCCAGTAATTGCAGGGGGTCGCTATTTGATTCAAGGAAAGTGGTATGTATTGATTTTTTTCTCACTTACTCTTTAGCATTTAGTTTAAA P. cer. Pipacs 1 ATAAAAGTTAGATTGTCATATGAAGAC-GTACACTCTTTCGATAAAACCTTGGGTGTTAGATAAATCATGATGTTGCTTTATCC-AACATAGGGGGAGGAGGGGTGGTTGTATGTTAGGA 238 P. cer. S-RNase-S36b2 ATAAAAGTTAGATTGTCATATGAAGAC-GTACACTCTTTCGATAAAACCTTGGGTGTTAGATAAATCATGATGTTGCTTTATCC-AACATAGGGGGAGGAGGGGTGGTTGTATGTTAGGA 238 P. cer. S36b3-RNase A-AAAAGTTAGATTGTCATATGAAGACAGTACACTCTTTCGATAAAACCTTGGGTGTTAGATAAATCCTGATGTTGCTTTATCCCAACATAGGGGGAGGAGGGGTGGTTGTATGTTAGGA 239 P. cer. S36b-RNase A-AAAAGTTAGATTGTCATATGAAGACAGTACACTCTTTCGATAAAACCTTGGGTGTTAGATAAATCCTGATGTTGCTTTATCCCAACATAGGGGGAGGAGGGGTGGTTGTATGTTAGGA 239 P. cer. S36a-RNase A-AAAAGTTAGATTGTCATATGAAGACAGTACACTCTTTCGATAAAACCTTGGGTGTTAGATAAATCCTGATGTTGCTTTATCCCAACATAGGGGGAGGAGGGGTGGTTGTATGTTAGGA P. cer. Pipacs 1 AATTCTCTCCACTCTTTTTTCTTTGATACAAGGGGAGGGGGAGGCGAGCCTAGGACATCGGGGGCATAGATAAATGCCTTTAACTACTTATTTGAAAGTCCTTTCCTTTTTAATTTAAGT 358 P. cer. S-RNase-S36b2 AATTCTCTCCACTCTTTTTTCTTTGATACAAGGGGAGGGGGAGGCGAGCCTAGGACATCGGGGGCATAGATAAATGCCTTTAACTACTTATTTGAAAGTCCTTTCCTTTTTAATTTAAGT 358 P. cer. S36b3-RNase AATTCTCTCCACTCTTTTTTCTTTGATACAAGGGGAGTGGGAGGCGAGCCTAGGACATCGGGTGCATAGATAAATGCCTTTAACTACTTATTTGAAAGTCTTTTCCTTTTTAATTTAAGT 359 P. cer. S36b-RNase AATTCTCTCCACTCTTTTTTCTTTGATACAAGGGGAGGGGGAGGTGAGCCTAGGACATCGGGTGCATAGATAAATGCCTTTAACTACTTATTTGAAAGTCCTTTCCTTTTTAATTTAAGT 359 P. cer. S36a-RNase AATTCTCTCCACTCTTTTTTCTTTGATACAAGGGGAGGGGGAGGTGAGCCTAGGACATCGGGTGCATAGATAAATGCCTTTAACTACTTATTTGAAAGTCCTTTCCTTTTTAATTTAAGT P. cer. Pipacs 1 TTTTCACTTTCATAATTTTATTTCATAAGATAAAAATTAAGTAATCAGTCCAGCTGTAAAAAATAATTTAATTTTTTTTAAGGTGGACACATGACCATATTTAAGTGGGAGTACCATAAT 478 P. cer. S-RNase-S36b2 TTTTCACTTTTATAATTTTATTTCATAAGATAAAAATTAAGTAATCAGTCCAGCTGTAAAAAATAATTTAATTTTTTTTAAGGTGGACACATGACCATATTTAAGTGGGAGTACCATTAT 478 P. cer. S36b3-RNase TTTTCACTTTTATAATTTTATTTCATAAGATAAAAATTAAGTTATTAGTCCAGCTGTAAAAAATAATTTAATTTTTTTTAAGGTGGACACATGACCATATTTAAGTGGGAGTACCATTAT 479 P. cer. S36b-RNase TTTTCACTTTTATAATTTTATTTCATAAGATAAAAATTAAGTAATTAGTCCAGCTGTAAAAAATAATTTTTTTTTTTTTAAGGTGGACACATGACCATATTTAAGTGGGAGTACCATTAT 479 P. cer. S36a-RNase TTTTCACTTTTATAATTTTATTTCATAAGATAAAAATTAAGTAATTAGTCCAGCTGTAAAAAATAAATTTTTTTTTTTTAAGGTGGACACATGACCATATTTAAGTGGGAGTACCATTAT P. cer. Pipacs 1 TGTTACATATTTTATTATTGTAAATCAAAATCACTAATTCAGTATAGTACTCAGGTTTAACGTAAAAATTATCTTATTCAAGAACGGAAATCTCTCT--TAAGTTTTTACTATTCCTTAA 596 P. cer. S-RNase-S36b2 TGTTACATATTTTATTATTGTAAATCAAAATCACTAATTCAGTATAGTACTCAGGTTTAACGTAAAAATTATCTTATTCAAGAACGGAAATCTCTCT--TAAGTTTTTACTATTCCTTAA 596 P. cer. S36b3-RNase TGTTACATATTTTATTATTGTAAATCAAAATCACTAATTCAGTATAGTACTCAGGTTTAACGTAAAAATTATCTTATTCAAGAACGGAAATCTCTCT--TAAGTTTTTACTATTCCTTAA 597 P. cer. S36b-RNase TGTTACATATTTTATTATTGTAAATCAAAATCACTAATTCAGTATAGTACTCAGGTTTAACGTAAAAATTATCTTATTCAAGAACGGAAATCTCTCTCTTAAGTTTTTACTATTCCTTAA 599 P. cer. S36a-RNase TGTTACATATTTTATTATTGTAAATCAAAATCACTAATTCAGTATAGTACTCAGGTTTAACGTAAAAATTATCTTATTCAAGAACGGAAATCTCTCTCTTAAGTTTTTACTATTCCTTAA P. cer. Pipacs 1 AATATGTATGAATTGCTTGGATGTCTCAGTACCCTCAGTTGCGATTGAATCTGAAGATATCTTGGCCCAACGTGAAAAGTGGCAATGATACAGAATTTTGGGAAAGCGAATGGAACAAA 715 P. cer. S-RNase-S36b2 AATATGTATGAATTGCTTGGATGTCTCAGTACCCTCAGTTGCGATTGAATCTGAAGATATCTTGGCCCAACGTGAAAAGTGGCAATGATACAGAATTTTGGGAAAGCGAATGGAACAAA 715 P. cer. S36b3-RNase AATATGTATGAATTGCTTGGATGTCTCAGTACCCTCAGTTGCGATTGAATCTGAAGATATCTTGGCCCAACGTGAAAAGTGGCAATGATACAGAATTTTGGGAAAGCGAATGGAACAAA 716 P. cer. S36b-RNase AATATGTATGAATTGCTTGGATGTCTCAGTACCCTCAGTTGCGATTGAATCTGAAGATATCTTGGCCCAACGTGAAAAGTGGCAATGATACAGAATTTTGGGAAAGCGAATGGAACAAA 718 P. cer. S36a-RNase AATATGTATGAATTGCTTGGATGTCTCAGTACCCTCAGTTGCGATTGAATCTGAAGATATCTTGGCCCAACGTGAAAAGTGGCAATGATACAGAATTTTGGGAAAGCGAATGGAACAAA 718 Confirmation from primers amplifying a 37-bp insert in the promoter region

13 Pipacs 1 might be an offspring of Cigánymeggy? Cultivars S-phenotype S-genotype* Érdi bőtermő SC S 4 S 6m S 35 S 36a Érdi jubileum SC S 1 S 6 S 13 S 36b Érdi nagygyümölcsű SI S 1 S 12 S 13 S 36b Meteor SC S 13m S 33 S 36a S 36b Montmorency SC S 6 S 13m S 35 S 36a Pándy clones SI S 1 S 4 S 35 S 36b Újfehértói fürtös SC S 19 S 4 S 35 S 36b Cigánymeggy SC S 6m2 S 9 S 26 S 36b2 Pipacs1 SC S 26 S 36b2 Former S-genotypes have been published in Tsukamoto et al. 21. Genetics 184: A63 Cigány Pipacs CP21 Cigány Pipacs B4 Cigány Pipacs CP12 Cigány Pipacs B2 Cigány Pipacs and so on

14 Relative gene expression Genes behind Amarelle-Morello phenotype PAL C4H CHS CHI F3H F3'H F3'5'H FLS DFR ANS 14 ANR 7 LAR 25 UFGT 8 MYB 7 MYB Ripening stages

15 SSR fingerprints of Érdi bőtermő clones

16 Potential new markers I. 2 bp 15 bp 1 bp 75 bp 5 bp Mk Úf Pá C4 Db ÉB N3 N5 L P1 julia.halasz@uni-corvinus.hu

17 Potential new markers I. Mb S-locus Distribution of the identified Prunus-specific miniature inverted repeat transposable elements, Falling Stones throughout the peach genome. Arrows indicate the orientation of the full length sequences and lines show fragmented copies. The approximate position of each centromere is indicated by an oval (not drawn to scale).

18 Potential new markers I. 1 bp 5 bp 25 bp M Genomic PCR analysis (in negative) of 18 different Prunus species and 4 species outside the Prunus genus using the FaSt-specific primers. M: 1 kb + DNA ladder, 1: P. armeniaca Armed-3 genotype; 2: P. dulcis Tétényi bőtermő ; 3: P. persica Babygold 7 ; 4: P. avium Germersdorfi óriás ; 5: P. cerasus Érdi jubileum ; 6: P. salicina Santa rosa ; 7: P. domestica Tophit ; 8: P. arabica; 9: P. tenella; 1: P. webbii; 11: P. armenica P. salicina hybrid; 12: P. kansuensis; 13: P. kurdica; 14: P. sogdiana; 15: P. mume; 16: P. ferganensis; 17: P. cerasifera; 18: P. spinosa; 19: Malus domestica Jonagold ; 2: Hippophae rhamnoides Frugana ; 21: Cydonia oblonga Aromate ; and 22: Cornus mas Császlói.

19 Future tasks To complete the S-genotypes and SSR fingerprints of the most perspective Hungarian sour cherry cultivars/selections To elucidate the putative origin of the most important Hungarian cultivars Sequencing MYB genes (and their promoter region) from amarelle- and morello-type cultivars Identification and application of transposable elements to develop new markers

20 Thank you for your kind attention! Dr. Júlia Halász Dr. Attila Hegedűs Dr. Sezai Ercişli Dr. Ossama Kodad Dr. Stefanovits-Bányai Éva Dr. Abrankó László Dr. Pfeiffer Péter Papp Nóra Dr. Tibor Szabó János Bolyai Scholarship, HAS Dr. József Nyéki Dr. Zoltán Szabó Ferenc Szőke OTKA K8429 NKTH-OTKA K68921 OTKA PD78124