Satsuma mandarin (Citrus unshiu Marcow.) chromosomes were stained with Giemsa and fluorochromes chromomycin A3 (CMA)/4′,6-diamidino-2-phenyindole (DAPI). Eighteen chromosomes were categorized into eight groups by the position and relative size of the CMA (+) region and relative length of chromosome. Ponkan (C. reticulata Blanco) DNA labeled with Dig-rhodamine (red) and pummelo [C. maxima (Burm.) Merr.] DNA labeled with biotin-fluorescein isothiocyanate (green) were used as genomic in situ hybridization (GISH) probes. GISH signals were detected on CMA (+) regions and other heterochromatin blocks. The chromosomes were categorized into 12 groups by the coloration and size of GISH signals with relative length of chromosomes. GISH allowed six pairs of speculated homozygous and six individual heterozygous chromosomes of satsuma mandarin to be identified unambiguously. In 10 chromosomes with distinct GISH signals on the CMA (+) regions, red GISH signals were detected on nine chromosomes, indicating that satsuma mandarin is closely related to ponkan. Two colors (red and green) of GISH signals were detected on type C chromosome and three different colors (red, green, and yellow) were detected on type A, indicating that pummelo is involved in the origin of satsuma mandarin. The origins of types A and C chromosomes in satsuma mandarin were also discussed. This article demonstrates that GISH is a powerful tool for chromosome identification and karyotyping in citrus.
Akira Kitajima, Atsu Yamasaki, Tsuyoshi Habu, Bannarat Preedasuttijit and Kojiro Hasegawa
Ryutaro Tao, Tsuyoshi Habu, Hisayo Yamane, Akira Sugiura and Kazuya Iwamoto
Self-compatible cultivars of Japanese apricot (Prunus mume Sieb. et Zucc.) have a horticultural advantage over self-incompatible ones because no pollinizer is required. Self-incompatibility is gametophytic, as in other Prunus species. We searched for molecular markers to identify self-compatible cultivars based on the information about S-ribonucleases (S-RNases) of other Prunus species. Total DNA isolated from five self-incompatible and six self-compatible cultivars were PCR-amplified by oligonucleotide primers designed from conserved regions of Prunus S-RNases. Self-compatible cultivars exhibited a common band of ≈1.5 kbp. Self-compatible cultivars also showed a common band of ≈12.1 kbp when genomic DNA digested with HindIII was probed with the cDNA encoding S 2-RNase of sweet cherry (Prunus avium L.). These results suggest that self-compatible cultivars of Japanese apricot have a common S-RNase allele that can be used as a molecular marker for self-compatibility.
Tsuyoshi Habu, Fumio Kishida, Miki Morikita, Akira Kitajima, Toshiaki Yamada and Ryutaro Tao
Japanese apricot (Prunus mume Sieb. et Zucc.) exhibits S-RNase-based gametophytic self-incompatibility as do other Prunus species. Both self-incompatible and self-compatible Japanese apricot cultivars are grown commercially in Japan. These self-compatible cultivars are shown to have a common S-haplotype called S f that contains S f-RNase and SFB f (S-haplotype-specific F-box protein). This study describes a simple and rapid detection of SFB f, in Japanese apricot, based on loop-mediated isothermal amplification (LAMP) method. A set of 4 primers, F3, B3, FIP, and BIP primer, were designed from the exon and the putative inserted sequence of SFB f. Optimal reaction time at 63 C was determined to be 90 minutes. It appeared that the LAMP method combined with the ultrasimple DNA extraction efficiently detected SFB f. The advantage of the marker-assisted selection of self-compatibility based on the LAMP method was discussed.