S-allele genotypes of nine apple (Malus ×domestica Borkh.) cultivars were identified using S-allele–specific polymerase chain reaction (PCR)–restriction fragmentlength polymorphism (RFLP) analysis. A new S-allele, Sg, was proposed to be present in `American Summer Pearmain', `Indo', `Kitanosachi', and `Meku 10'. This allele is very similar to Sf at the nucleotide sequence (92%) and deduced amino acid sequence (94%) levels.
Seasonal changes in the amounts of the NAD-dependent sorbitol dehydrogenase (NAD-SDH) (enzyme code, 188.8.131.52) protein in developing apple (Malus pumila Mill var. domestica Schneid) fruit were determined by immunoblotting analysis. The amounts of the enzyme protein were very low in young fruit and rose as fruit matured. The weak correlation between enzyme protein and NAD-SDH activity and also the changes in NAD-SDH specific activity suggested that there could be posttranslational modification to the pre-existing enzyme or isoenzyme(s) of NAD-SDH. The changes in the amounts of NAD-SDH protein did not show the same pattern as those in relative growth rate, which is used to express sink activity, especially in young fruit. The role of NAD-SDH on sink activity in apple fruit, therefore, could not be explained simply by the amount and activity of the enzyme. In young fruit, it seems that enzymes other than NAD-SDH would be more directly related with fruit growth.
The S-locus genes in the pistil (S-RNases) were cloned from the apple (Malus ×domestica Borkh.) cultivar Akane (S-genotype SdSh from pollination analysis). The Sd- and Sh-RNase corresponded to S7- and S24-RNase, which have been cloned from `Idared' and `Braeburn', respectively. Sh-RNase was very similar to Sf- and Sg-RNases at the deduced amino acid-sequence levels (93%). We developed an S-allele specific polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis method for distinguishing the Sh from Sf and Sg, and the Sh-alleles of `Akane', `Touhoku 2', `Vista Bella', and `Worcester Pearmain' were identified. We also identified the S-allele genotypes of 16 apple cultivars.
The firmness of the flesh in 27 apple (Malus ×domestica Borkh.) cultivars and selections (genotypes) was measured as an indicator of storage potential at 20 days after harvest under 20 ± 2 °C, 80% ± 5%relative humidity storage conditions. Softening ranged from 9% to 58% of initial values among genotypes after 20 days of storage. In some genotypes, softening was not continuous, a minimum firmness being reached before day 20. After a period of rapid softening, firmness declined to at least 20% of that at harvest. For each genotype, linear regression analysis of firmness changes from harvest until when firmness decreased by 20% was carried out. In genotypes in which firmness did not drop >20% within 20 days of storage, the entire dates to 20 days were used for analysis. The homogeneity of the regression residual variances and their normal distribution was not rejected at P = 0.05, and the linear regression analysis was assumed to be applicable to the change in firmness for each genotype. Results of the regression analysis showed that the regression was significant for all genotypes except one. Therefore, storage potential could be evaluated by comparing the regression coefficient of each genotype.
Aqueous ethanol extracts prepared from 19 apple (Malus ×domestica Borkh.) cultivars were studied to explore their antiproliferative activity. Half of them showed strong inhibition on proliferation of human leukemic HL-60 cells, while the others were weak. Total polyphenols, 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, and total anthocyanins were measured and the results indicated that the antiproliferative activity was more strongly correlated to the polyphenols and radical scavenging activity than to the anthocyanin content. Several polyphenols in `Jonathan' were identified and quantified by high-performance liquid chromatography (HPLC) analysis. Among those compounds found during HPLC, catechin and epicatechin seemed partially responsible for HL-60 antiproliferation. A careful examination on parentage of the apple cultivars tested revealed that `Jonathan' and its progeny showed high antiproliferation toward HL-60. This is the first observation about the relationship between antiproliferative activity and parentage of apples, and the information would be useful to create new apple cultivars that posses more anticancer potential.
As the parents of the some of the apple cultivars were unknown and others were uncertain, we investigated the parent-offspring relationships of eight apple cultivars by S-RNase analysis and SSR markers. The paternal parent of `Hida' was identified as `Golden Delicious', not the previously mentioned `Orin'. It was indicated that `Ryoka No Kisetsu' and `Korin' showing identical SSR genotype are likely sports of `Fuji'. `Fuji', rather than `Toko', seemed to be a maternal parent of `Kotoku', but was not a paternal parent of `Orei', `Starking Delicious', `Nero 26', `Empire', or `Aori 3'. Previously mentioned `Mutsu', `Indo', and `Shin Indo' were excluded as paternal parents of `Hokuto'. `Tsugaru' and `Jonathan' and were identified as the respective paternal parents of three cultivars described as having unknown paternal parents, i.e., `Aika No Kaori', `Yoko', and `Tsugaru'.
We examined the genetic diversity and relatedness among apple (Malus ×domestica Borkh.) cultivars in Japan. The 42 apple cultivars, including major cultivars in Japan, were divided into five groups based on SSR genotypes. Most economically important cultivars belong in three groups: Fuji-Delicious, Golden Delicious, and Jonathan groups, and their genetic backgrounds seemed to be narrow. We also investigated the parent-offspring relationships of nine apple cultivars. `Jonathan', `Fuji', and `Rero 11' were identified as the respective paternal parents of three cultivars described as having unknown paternal parents (i.e., `Akagi', `Ambitious', and `Hokuto'). `Starking Delicious', `Senshu', and `Golden Delicious', rather than `Ralls Janet', `Hatsuaki', and `Indo', seemed to be the paternal parents of `Kinsei', `Kiou', and `Mellow', respectively. `Carolina Red June' was excluded as a paternal parent of `Ranzan'. Both attributed parents of `Scarlet' (`Akane' and `Starking Delicious') were excluded, and it was suggested that `Fuji' was used as either a maternal or a paternal parent of `Scarlet'. `Jonathan' rather than `McIntosh' seems to be a maternal parent of `Yukari'.
Two apple [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.] homologous fragments of FLO/LFY and SQUA/AP1 (AFL and MdAP1, respectively) were analyzed to determine the relationship between floral bud formation and floral gene expression in `Jonathan' apple. The AFL gene was expressed in reproductive and vegetative organs. By contrast, the MdAP1 gene, identified as MdMADS5, which is classified into the AP1 group, was expressed specifically in sepals concurrent with sepal formation. Based on these results, AFL may be involved in floral induction to a greater degree than MdAP1 since AFL transcription increased ≈2 months earlier than MdAP1. Characterization of AFL and MdAP1 should advance the understanding of the processes of floral initiation and flower development in woody plants, especially in fruit trees like apple.