Screening for resistance to Nectria galligena Bres. in cut shoots of apple

Summary To find a fast and reliable test to assess resistance to Nectria galligena in apple, different methods of inoculation were compared using macroconidia of N. galligena and one-year-old cut shoots from mature trees of Cox's Orange Pippin, IVT 69078-19, James Grieve and Jonathan.With the best inoculation method 11 genotypes were screened for resistance. Elstar, Golden Delicious, Jonathan and Lombart's Calville were highly resistant and the level of resistance of Ingrid marie, Gloster, Melrose, IVT 69078-19, Cox's Orange Pippin, James Grieve and Idared decreased in this order.The best inoculation method proved to be simple, giving results within four to nine weeks after inoculation.     

Investigation and genetic mapping of a Glomerella leaf spot resistance locus in apple

Apple Glomerella leaf spot (GLS) is a severe fungal disease that damages apple leaves during the summer in China. Breeding new apple varieties that are resistant to the disease is considered the best way of controlling GLS. Fine mapping and tightly linked marker are critically essential for the preselection of resistant seedlings. In this study, a population of 207 F₁ individuals derived from a cross between ‘Golden Delicious’ and ‘Fuji’ was used to construct a fine simple sequence repeat (SSR)‐based genetic linkage map. The position of R_gls, a locus responsible for resistance to GLS, was identified on apple linkage group (LG) 15 using SSR markers CH05g05 and CH01d08, which was adapted from a published set of 300 SSR markers that were developed using the bulked segregant analysis (BSA) method. These two SSR markers flanked the gene, and its recombination rate was 8.7% and 23.2%, respectively. A total of 276 newly developed SSR markers around the target region and designed from the genome apple assembly contig of LG15 were screened. Only nine of these were determined to be linked to the R_gls locus. Thus, a total of 11 SSR markers were in linkage with R_gls, and mapped at distances ranging from 0.5 to 33.8 cM. The closest marker to the R_gls locus was S0405127, which showed a genetic distance of approximately 0.5 cM. The first mapping of the gene R_gls was constructed, and the locations of the 11 effective primers in the ‘Golden Delicious’ apple genome sequence were anchored. This result facilitates better understanding of the molecular mechanisms underlying the trait of resistance to GLS and could be used in improving the breeding efficiency of GLS‐resistant apple varieties.     

Effects of preharvest nitric oxide treatment on ethylene biosynthesis and soluble sugars metabolism in ‘Golden Delicious’ apples

In order to examine the influence of preharvest nitric oxide (NO) treatment on ethylene biosynthesis and soluble sugar metabolism in ‘Golden Delicious’ apples, apple trees were sprayed with 50 μM sodium nitroprusside (SNP) (a donor of NO) 14 days before harvest. The results indicated that preharvest SNP treatment can increase the NO content and the NOS activity in apple fruit, therefore, delay the accumulation of ethylene due to its inhibition on the activities of 1-aminocyclopropane-1-carboxylate synthase (ACS) and 1-aminocyclopropane-1-carboxylate oxydase (ACO). Fructose is the main sugar in ‘Golden Delicious’ apple. The synthesis of sucrose was stimulated and the decomposition of sucrose was inhibited by this treatment, thus causing the accumulation of sucrose. We can draw a conclusion that pre-harvest SNP (50 μM) treatment can increase the NO content of fruit during storage, while higher NO content can further regulate fruit ripening through its effect on ethylene and sugar metabolism in ‘Golden Delicious’ apple fruit during storage at 18 °C.     

Effects of auxin and jasmonates on 1-aminocyclopropane-1-carboxylate (ACC) synthase and ACC oxidase gene expression during ripening of apple fruit

1-Aminocyclopropane-1-carboxylate (ACC) synthase and oxidase activities, their gene expression, and ethylene production in apple fruit [Malus sylvestris (L.) Mill. Var. domestica (Borkh.) Mansf.] treated with a synthetic auxin 2,4-dichlorophenoxy-propionic acid (2,4-DP) and n-propyl dihydrojasmonate (PDJ), a jasmonic acid derivative, has been investigated to clarify the action of auxin and jasmonates on ethylene production. The fruit was harvested at 103 d after full bloom (preclimacteric). The expression of MdACS4 messenger RNA (mRNA) at 48 and 96 h after treatment was higher in fruit treated with 2,4-DP than in the untreated control, but those of MdACS1 and MdACO1 were not affected by treatment. The ethylene production in 2,4-DP-treated fruit increased at 96 h after treatment. In contrast, expression of mRNAs hybridized with MdACS1 and MdACO1 probes in the skin of PDJ-treated fruit were higher than those in the untreated control. In addition, ACC synthase activity and ethylene production also increased after treatment. These results show that the ethylene production rate may differ with the kind of genes which were stimulated by auxin or jasmonates.     

A new strain of Metschnikowia fructicola for postharvest control of Penicillium expansum and patulin accumulation on four cultivars of apple

The efficacy of three antagonistic yeasts, Metschnikowia pulcherrima strain MACH1, M. pulcherrima strain GS9, and Metschnikowia fructicola strain AL27, against Penicillium expansum and patulin accumulation was evaluated on apples stored at room (22 ± 1 °C for 7 days) and cold temperatures (1 ± 1 °C for 56 days). To increase the potential range of application of the biocontrol agents (BCAs), their efficacy was evaluated on four cultivars of apple, i.e. ‘Golden Delicious’, ‘Granny Smith’, ‘Red Chief’ and ‘Royal Gala’. AL27 was more effective than MACH1 and GS9 in the control of blue mold rot and in the reduction of patulin accumulation. The efficacy of AL27 was in most cases similar to the chemical control used, making the antagonist as competitive as chemical fungicides. In vitro experiments showed that AL27 reduced the conidial germination and germ tube length of P. expansum more than the other strains. The three BCAs were more effective in the control of blue mold rot on ‘Golden Delicious’ apples than on the other tested cultivars.     

Identification and validation of an over‐dominant QTL controlling soybean seed weight using populations derived from Glycine max × Glycine soja

Heterosis, or hybrid vigour, has been used to improve seed yield in several important crops for decades and it has potential applications in soybean. The discovery of over‐dominant quantitative trait loci (QTL) underlying yield‐related traits, such as seed weight, will facilitate hybrid soybean breeding via marker‐assisted selection. In this study, F₂ and F_2 : 3 populations derived from the crosses of ‘Jidou 12’ (Glycine max) × ‘ZYD2738’ (Glycine soja) and ‘Jidou 9’ (G. max) × ‘ZYD2738’ were used to identify over‐dominant QTL associated with seed weight. A total of seven QTL were identified. Among them, qSWT_13_1, mapped on chromosome 13 and linked with Satt114, showed an over‐dominant effect in two populations for two successive generations. This over‐dominant effect was further examined by six subpopulations derived from ‘Jidou12’ × ‘ZYD2738’. The seed weight for heterozygous individuals was 1.1‐ to 1.6‐fold higher than that of homozygous individuals among the six validation populations examined in different locations and years. Therefore, qSWT_13_1 may be a useful locus to improve the yield of hybrid soybean and to understand the molecular mechanism of heterosis in soybean.     

Genotype analysis of the wheat semidwarf Rht‐B1b and Rht‐D1b ancestral lineage

In wheat, semidwarfism resulting from reduced height (Rht)‐B1b and Rht‐D1b was integral to the ‘green revolution’. The principal donors of these alleles are ‘Norin 10’, ‘Seu Seun 27’ and ‘Suwon 92’ that, according to historical records, inherited semidwarfism from the Japanese landrace ‘Daruma’. The objective of this study was to examine the origins of Rht‐B1b and Rht‐D1b by growing multiple seed bank sources of cultivars comprising the historical pedigrees of the principal donor lines and scoring Rht‐1 genotype and plant height. This revealed that ‘Norin 10’ and ‘Suwon 92’ sources contained Rht‐B1b and Rht‐D1b, but the ‘Seu Seun 27’ source did not contain a semidwarf allele. Neither Rht‐B1b nor Rht‐D1b could be definitively traced back to ‘Daruma’, and both ‘Daruma’ sources contained only Rht‐B1b. However, ‘Daruma’ remains the most likely donor of Rht‐B1b and Rht‐D1b. We suggest that the disparity between historical pedigrees and Rht‐1 genotypes occurs because the genetic make‐up of seed bank sources differs from that of the cultivars actually used in the pedigrees. Some evidence also suggests that an alternative Rht‐D1b donor may exist.     

Inhibition of browning on the surface of apple slices by short term exposure to nitric oxide (NO) gas

Freshly cut slices of apple (Malus x domestica Borkh cv. Granny Smith) were fumigated with nitric oxide (NO) gas at concentrations between 1 and 500 μl l⁻¹ in air at 20 °C for up to 6 h followed by storage at 0, 5, 10 and 20 °C in air. Exposure to nitric oxide delayed the onset of browning on the apple surface with the most effective treatment being fumigation with 10 μl l⁻¹ NO for 1 h. While nitric oxide inhibited browning in slices held at all temperatures, it was relatively more effective as the storage temperature was reduced with the extension in postharvest life over the respective untreated slices increasing from about 40% at 20 °C to about 70% at 0 °C. In a smaller study on ‘Royal Gala’, ‘Golden Delicious’, ‘Sundowner’, ‘Fuji’ and ‘Red Delicious’ slices stored at 10 °C, 10 μl l⁻¹ NO for 1 h was found to be effective in inhibiting surface browning in all cultivars.     

Characterization of broad‐spectrum resistance to Soybean mosaic virus in soybean [Glycine max (L.) Merr.] cultivar ‘RN‐9’

Soybean mosaic virus (SMV) can cause serious yield losses in soybean. Soybean cultivar ‘RN‐9’ is resistant to 15 of 21 SMV strains. To well‐characterize this invaluable broad‐spectrum SMV‐resistance, populations (F₁, F₂ and F_2:3) derived from resistant (R) × susceptible (S) and R × R crosses were tested for SMV‐SC18 resistance. Genetic analysis revealed that SC18 resistance in ‘RN‐9’ plus two elite SMV‐resistant genotypes (‘Qihuang No.1’ and ‘Kefeng No.1’) are controlled by independently single dominant genes. Linkage analysis showed that the resistance of ‘RN‐9’ to SMV strains SC10, SC14, SC15 and SC18 is controlled by more than one gene(s). Moreover, Rsc10‐r and Rsc18‐r were both positioned between the two simple sequence repeats markers Satt286 and Satt277, while Rsc14‐r was fine‐mapped in 136.8‐kb genomic region containing sixteen genes, flanked by BARCSOYSSR_06_0786 and BARCSOYSSR_06_0790 at genetic distances of 3.79 and 4.14 cM, respectively. Allelic sequence comparison showed that Cytochrome P450‐encoding genes (Glyma.06g176000 and Glyma.06g176100) likely confer the resistance to SC14 in ‘RN‐9’. Our results would facilitate the breeding of broad‐spectrum and durable SMV resistance in soybeans.     

Validation of a male‐linked gene locus (OGI) for sex identification in persimmon (Diospyros kaki Thunb.) and its application in F1 progeny

It is crucial to develop a rapid technique for identifying sexuality in the seedling stage of persimmon (Diospyros kaki Thunb.), and the elimination of male progeny has been regarded as an important strategy for enhancing breeding efficiency. In this study, phenotype characterization and genotyping of the male‐linked OGI marker were carried out using 205 accessions, including persimmon cultivars, F₁ progeny and nine related Diospyros species. All persimmon cultivars displayed consistent results regarding OGI amplification and sex phenotype. A total of 143 F₁ progeny were derived from 11 crosses, among which 95 individuals had flowered. In the flowering full‐sib families, the amplification of the OGI marker in agreement with the sex phenotype was obtained in 85 plants (89.5%). The segregation of OGI in ‘Huashi 1’ × ‘Luotian Tianshi’ and ‘Huashi 1’ × Male 3 F₁ populations fit a 1 : 1 ratio. Furthermore, high OGI transferability was observed in nine related species. Overall, the results indicated that the OGI locus could be used to distinguish male from female persimmon plants at an early stage.     

Mapping seedling resistance to net form of net blotch (Pyrenophora teres f. teres) in barley using detached leaf assay

To develop molecular markers against Pyrenophora teres f. teres in barley, a detached leaf assay was conducted on two DH populations with a set of 11 single conidial lines (SCLs). Out of these, three showed different reactions in the DH population ‘Uschi × HHOR3073’ and two in ‘(P x V) × HHOR9484’. For SCL ‘QLB’, a 1r : 1s (χ² = 2.78) segregation was observed in the population ‘Uschi × HHOR3073’. In contrast to this, a continuous variation was observed for the SCL ‘WvB’ and ‘d8_4’ in the DH population ‘Uschi × HHOR3073’ and for ‘AR’ and ‘net1840’ in ‘(P × V) × HHOR9484’. With respect to resistance to the SCL ‘QLB’, a single major gene was located on chromosome 7H, and for resistance against SCL ‘WvB’, two QTLs were detected on chromosome 3H and 7H, and against SCL ‘d8_4’, two loci were mapped on chromosome 3HS in ‘Uschi × HHOR3073’. In the DH population ‘(P x V) × HHOR9484’, one locus conferring resistance to the SCL ‘AR’ was located on chromosome 3H. For resistance to SCL ‘net1840’, two QTLs were mapped on chromosome 4H and 5H.     

A PCR-based DNA marker for detection of mutant and normal alleles of the Wx-D1 gene of wheat

To assist waxy wheat breeding a DNA marker was developed to discriminate mutant and normal alleles at the Wx‐D1 locus. This polymerase chain reaction‐based marker distinguishes the mutant from the normal allele by targeting the previously reported deletion basis of the mutant. The marker codominantly identifies the normal allele of the Wx‐D1 gene from the mutant allele originated from the Chinese landrace ‘Baihoumai’. However, attempts with a number of primer combinations targeting this deletion failed to amplify the corresponding fragment from an unrelated wheat line (NP150) that has a mutant null allele at the same locus. This indicates that NP150 has a different mutant allele from that of ‘Baihoumai’. This marker is a useful tool to identify wheat cultivars with mutant and normal alleles of the Wx‐D1 gene, and is used in marker‐assisted selection of the Wx‐D1 gene in our waxy wheat breeding programme.     

Evaluation of cold hardiness in two sets of near-isogenic lines of wheat (Triticum aestivum) with polymorphic vernalization alleles

In wheat, variation at the orthologus Vrn‐1 loci, located on each of the three genomes, A, B and D, is responsible for vernalization response. A dominant Vrn‐1a allele on any of the three wheat genomes results in spring habit and the presence of recessive Vrn‐1b alleles on all three genomes results in winter habit. Two sets of near‐isogenic lines (NILs) were evaluated for DNA polymorphisms at their Vrn‐A1, B1 and D1 loci and for cold hardiness. Two winter wheat cultivars, ‘Daws’ and ‘Wanser’ were used as recurrent parents and ‘Triple Dirk’ NILs were used as donor parents for orthologous Vrn‐1 alleles. The NILs were analysed using molecular markers specific for each allele. Only 26 of 32 ‘Daws’ NILs and 23 of 32 ‘Wanser’ NILs had a plant growth habit that corresponded to the marker genotype for the markers used. Freezing tests were conducted in growth chambers programmed to cool to ?21.5°C. Relative area under the death progress curve (AUDPC), with a maximum value of 100 was used as a measure of death due to freezing. The average relative AUDPC of the spring habit ‘Daws’Vrn‐A1a NILs was 86.15; significantly greater than the corresponding winter habit ‘Daws’Vrn‐A1b NILs (42.98). In contrast, all the ‘Daws’Vrn‐A1bVrn‐B1aVrn‐D1b and Vrn‐A1bVrn‐B1bVrn‐D1a NILs (spring habit) had relative AUDPC values equal to those of their ‘Daws’ sister genotypes with Vrn‐A1bVrn‐B1bVrn‐D1b NILs (winter habit). The average AUDPC of spring and winter habit ‘Wanser’ NILs differed at all three Vrn‐A1, Vrn‐B1 and Vrn‐D1 locus comparisons. We conclude that ‘Daws’ and ‘Wanser’ have different background genetic interactions with the Vrn‐1 loci influencing cold hardiness. The marker for Vrn‐A1 is diagnostic for growth habit and cold hardiness but there is no relationship between the Vrn‐B1 and Vrn‐D1 markers and the cold tolerance of the NILs used in this study.     

Effect of the ph1b mutant on chromosome pairing in hybrids between Dasypyrum villosum and Triticum aestivum

Chromosome pairing was analysed in F₁ hybrids of the wheat cultivar ‘Chinese Spring’ (CS) and its ph1b mutant (CSphlb) with Dasypyrum villosum. On average, 1.61 chromosomes per cell paired in the hybrid CS ×D. villosum, but 14.43 in the hybrid CS ph1b×D. villosum. Genomic fluorescence in situ hybridization (GISH) revealed three types of homoeologous association between wheat (W) and D. villosum (D) chromosomes (W‐D, D‐W‐W and D‐W‐D) in pollen mother cells of the CS ph1b×D. villosum hybrid, and only one type (W‐W), in the CS ×D. villosum hybrid. Both F₁ hybrids were self‐sterile. The seed set of the backcross of CS ×D. villosum with CS was 6.67% and that of CS ph1b×D. villosum with CS or CS ph1b was only 0.45%. The chromosome number of BC₁ plants varied from 48 to 72. Translocations of chromosome segments or entire arms between wheat and D. villosum chromosomes were detected by GISH in the BC₁ plants from the backcross of CS ph1b×D. villosum to CS ph1b.     

Identification and molecular mapping of Flowering Date1 (FD1), a major photoperiod insensitivity gene in the adzuki bean (Vigna angularis)

The induction of flowering under long‐day conditions is an important adaptation by short‐day plants, such as adzuki beans (Vigna angularis), to high‐latitude environments. This study clarified the genetic control underlying the long‐day insensitivity of adzuki bean cultivar ‘Shumari’. ‘Shumari’ was found to be insensitive to a 16‐h day, whereas landrace Acc2265 was highly sensitive. When grown under natural long‐day conditions at Obihiro (42°9′N), Acc2265 initiated flowering at least 80 days after ‘Shumari’. When 86 recombinant inbred lines (RILs) derived from crosses between ‘Shumari’ and Acc2265 were grown under these conditions, their flowering dates ranged from the middle of July to the end of October. The distinct bimodal distribution in the RIL population was due to a single major gene, designated Flowering Date1 (FD1). Molecular mapping showed that FD1 was located between the SSR markers Az02‐37M3 and Az02‐40M9, at distances of 6 and 10.4 cM, respectively, on linkage group 2. RILs carrying FD1^S lacked long‐day sensitivity, whereas RILs carrying FD1^A were sensitive to long‐day conditions, confirming that FD1 controls long‐day sensitivity.