Summary• Breeding of apple (Malus · domestica) remains a slow process because of protracted generation cycles. Shortening the juvenile phase to achieve the introgression of traits from wild species into prebreeding material within a reasonable time frame is a great challenge.• In this study, we evaluated early flowering transgenic apple lines overexpressing the BpMADS4 gene of silver birch with regard to tree morphology in glasshouse conditions. Based on the results obtained, line T1190 was selected for further analysis and application to fast breeding.• The DNA sequences flanking the T-DNA were isolated and the T-DNA integration site was mapped on linkage group 4. The inheritance and correctness of the T-DNA integration were confirmed after meiosis. A crossbred breeding programme was initiated by crossing T1190 with the fire blight-resistant wild species Malus fusca. Transgenic early flowering F 1 seedlings were selected and backcrossed with 'Regia' and 98 ⁄ 6-10 in order to introgress the apple scab Rvi2, Rvi4 and powdery mildew Pl-1, Pl-2 resistance genes and the fire blight resistance quantitative trait locus FB-F7 present in 'Regia'.• Three transgenic BC'1 seedlings pyramiding Rvi2, Rvi4 and FB-F7, as well as three other BC'1 seedlings combining Pl-1 and Pl-2, were identified. Thus, the first transgenic early flowering-based apple breeding programme combined with marker-assisted selection was established.

Fire blight (FB), caused by the Gram-negative bacterium Erwinia amylovora is a dangerous disease on pome fruit, including apple. The FB-susceptible cultivar ÔIdaredÕ was crossed with the resistant wild species clone Malus · robusta 5. A segregating population of 146 progeny has been tested by artificial shoot inoculation for susceptibility to FB. Progeny were infected from 0% to 100% of the shoot length. To identify chromosomal regions or loci responsible for resistance to FB of Malus · robusta 5, a set of microsatellite markers (simple sequence repeat, SSRs) was chosen covering all linkage groups of apple. Up to eight different microsatellites were bulked to one mutliplex PCR using four different labels and a fifth label for a size standard. Fifty-nine microsatellite markers out of 72 SSRs were polymorphic. Fifty-four of 66 loci detected could be mapped and were useful for the detection of related resistant loci. Alleles of microsatellites Hi03d06, CH03g07 and CH03e03 originating from the resistant donor M. robusta were associated with resistance to Erwinia amylovora. Up to eighty percent of the phenotypic variation could be explained by the interval spanned by SSRs CH03g07 and CH03e03, indicating the presence of a major resistance gene. All three microsatellites are located on the distal part of linkage group 3, spanning 15 cM. The SSR marker CH03e03 can be regarded as diagnostic marker for FB resistance. Only seven progeny expressing allele b (184 bp) of CH03e03 showed blighted shoot lengths of more than 30% and only nine progeny lacking allele b showed blighted shoot lengths of <30%. By setting a threshold of 30% shoot necrosis for resistance to FB, the 146 individuals segregate into 71 susceptible and 75 resistant plants, and resistance to FB maps 9 cM away from marker CH03e03.

To shorten the juvenile stage of apple (Malus · domestica Borkh.) the BpMADS4 gene from silver birch (Betula pendula Roth.) was constitutively overexpressed in 25 transgenic apple clones. All clones were characterized by PCR, RT-PCR and Real Time PCR. Solitary flowers were produced on in vitro shoots of eight transgenic clones and most of them appeared to be morphologically normal. Twenty shoots of each clone were rooted and transferred to a glasshouse. Glasshouse plants of clones T1165, T1187 and T1190 developed flowers. Several plants of T1165 and T1187 started floral initiation within 3-4 months following transfer to the glasshouse. Primary flowers were solitary and in a terminal position on the main shoot. Lateral flower clusters, consisting of three to five individual flowers, were also found. Pollen vitality and tube germination of glasshouse-grown flowers were investigated, and there were no significant differences compared to pollen of nontransgenic control plants. Preliminary crosses using flowers of glasshouse plants resulted in small apple fruits. It would seem that this is the first report on in vitro flower induction in transgenic apple.

SummaryThe effects of daylength and temperature on flowering of the cultivated octoploid strawberry (Fragaria × ananassa Duch.) have been studied extensively at the physiological level, but information on the molecular pathways controlling flowering in the species is scarce. The flowering pathway has been studied at the molecular level in the diploid short‐day woodland strawberry (F. vesca L.), in which the FLOWERING LOCUS T1 (FvFT1)–SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (FvSOC1)–TERMINAL FLOWER1 (FvTFL1) pathway is essential for the correct timing of flowering. In this work, we show by transgenic approach that the silencing of the floral repressor FaTFL1 in the octoploid short‐day cultivar ‘Elsanta’ is sufficient to induce perpetual flowering under long days without direct changes in vegetative reproduction. We also demonstrate that although the genes FaFT1 and FaSOC1 show similar expression patterns in different cultivars, the regulation of FaTFL1 varies widely from cultivar to cultivar and is correlated with floral induction, indicating that the transcription of FaTFL1 occurs at least partially independently of the FaFT1–FaSOC1 module. Our results indicate that changing the expression patterns of FaTFL1 through biotechnological or conventional breeding approaches could result in strawberries with specific flowering and runnering characteristics including new types of everbearing cultivars.

SummaryFire blight is a destructive bacterial disease caused by Erwinia amylovora affecting plants in the family Rosaceae, including apple. Host resistance to fire blight is present mainly in accessions of Malus spp. and is thought to be quantitative in this pathosystem.In this study we analyzed the importance of the E. amylovora effector avrRpt2 EA , a homolog of Pseudomonas syringae avrRpt2, for resistance of Malus 9 robusta 5 (Mr5).The deletion mutant E. amylovora Ea1189DavrRpt2 EA was able to overcome the fire blight resistance of Mr5. One single nucleotide polymorphism (SNP), resulting in an exchange of cysteine to serine in the encoded protein, was detected in avrRpt2 EA of several Erwinia strains differing in virulence to Mr5. E. amylovora strains encoding serine (S-allele) were able to overcome resistance of Mr5, whereas strains encoding cysteine (C-allele) were not. Allele specificity was also observed in a coexpression assay with Arabidopsis thaliana RIN4 in Nicotiana benthamiana. A homolog of RIN4 has been detected and isolated in Mr5.These results suggest a system similar to the interaction of RPS2 from A. thaliana and AvrRpt2 from P. syringae with RIN4 as guard. Our data are suggestive of a gene-for-gene relationship for the host-pathogen system Mr5 and E. amylovora.