_??_1993 The Japan Mendel Society Cytologia 58: 313 -320 , 1993 Karyotypes on Five Species of Japanese Geum (Rosaceae) Yoshikane Iwatsubo and Naohiro Naruhashi Department of Biology, Faculty of Science, Toyama University, Gofuku, Toyama 930, Japan Accepted May 28, 1993 Geum Linn., a genus of the Rosaceae, comprises about 40 species occurring in the tem perate and arctic regions of both hemispheres (Airy Shaw 1973). Many cytological studies clarified that the species consist of a polyploid series with x=7 and 2n=28, 42, 56, 70 or 84 (Fedorov 1969). While the chromosome numbers for most species in the genus have been determined, no karyotype analyses have been made. In Japan there are five species of Geum; G. aleppicum, G. calthaefolium var. nipponicum, G. japonicum, G. macrophyllum var. sacha linense and G. pentapetalum (Ohwi 1965). Geum calthaefolium var. nipponicum is sometimes placed in Acomastylis, while G. pentapetalum is treated as Sieversia, by some workers (Bolle 1933, Hara 1936, Okuyama 1977, Shimizu 1982, Sugimoto 1978). This paper intends to clarify the cytological relationship of the five Japanese species of Geum through karyotypic studies. Materials and methods The five Japanese species of Geum which were studied are listed in Table 1, along with their collection localities. These plants were grown in the botanic garden of Toyama Uni versity. Root tips were collected from the potted plants, pretreated in a 2mM 8-hydroxyqu inoline solution for one hour at room temperature and then for 15 hr at 5•Ž. After being fixed in a mixture of glacial acetic acid and absolute ethyl alcohol (1:3) for one hour, the root tips were soaked in 1N HCl for a few hours, macerated in 1N HCl at 60•Ž for 11.5min, and immersed in tap water. Root tips were stained and squashed in 1.5% lacto-propionic orcein. Karyotypes were analyzed at mitotic metaphase. Chromosome form was expressed according to the nomenclature of Levan et al. (1964). Results (1) Geum aleppicum Jacq. Two plants collected from the same locality were studied (Table 1). Both had 2n=42 chromosomes (Fig. 1A), which confirmed previous reports (Yamazaki 1936, Hara 1952, Gajewski 1948, 1957). The karyotype was examined in one plant (Figs. 2A, 3A). Meta phase chromosomes ranged from 0.8ƒÊm-2.0ƒÊm in length and 1.0 to 4.0 in arm ratio (Table 2). The 42 chromosomes were classified into three groups: 12 metacentric pairs, seven sub metacentric pairs, and two subtelocentric pairs. The two submetacentric pairs had satellites on the short arms. The karyotype was formulated as follows: 2n=42=24m+10sm+4tsm +4st. (2) Geum calthaefolium Smith var. nipponicum (F. Bolle) Ohwi Ten plants from the same locality were examined (Table 1). All had 2n=42 chromosomes (Fig. 1B), which was consistent with the report, under the name of Acomastylis calthifolium 314 Yoshikane Iwatsubo and Naohiro Naruhashi Cytologia 58 (Smith) Bolle, of Hara (1952). Karyotypes in the two plants studied were nearly the same. Metaphase chromosomes ranged from 0.7ƒÊm-1.8ƒÊm in length and 1.0 to 6.0 in arm ratio (Table 3). They were classified into three groups: 16 metacentric pairs, three submetacentric pairs, two subtelocentric pairs (Figs. 2B, 3B). The longer subtelocentric pair had a satellite Fig. 1. Somatic metaphase chromosomes of five Japanese species of Geum. A: G. aleppicum, 2n=42; B: G. calthaefolium var. nipponicum, 2n=42; C: G. japonicum, 2n=42; D: G. macrophyllum var. sachalinense, 2n=42; E: G. pentapetalum, 2n=14. Bar represents 10ƒÊm. Table 1. Chromosome number, and collection locality or source of studied five Japanese species in Geum 1993 Karyotypes on Five Species of Japanese Geum 315 Table 2. Measurements of somatic metaphase chromosomes of Geum aleppicum t: satellite. Table 3. Measurements of somatic metaphase chromosomes of Geum calthaefolium var. nipponicum t: satellite. 316 Yoshikane Iwatsubo and Naohiro Naruhashi Cytologia 58 on the short arm. The karyotype of this plant was thus formulated as 2n=42=32m+6sm+ 2st+2tst. (3) Geum japonicum Thunb. ex Murray Four plants, each from a different locality, were examined (Table 1). All plants had 2n=42 (Fig. 1C), which confirmed the previous reports (Yamazaki 1936, Gajewski 1949, 1957). Karyotypes in the three plants studied showed no marked differences. Metaphase chromo somes ranged from 0.8ƒÊm-2.1ƒÊm in length and 1.1 to 4.0 in arm ratio (Table 4). Somatic Fig. 2. Karyotypes of five Japanese species of Geum. A: G. aleppicum, 2n=42; B: G. calthaefolium var. nipponicum, 2n=42; C: G . japonicum, 2n= 42; D: G. macrophyllum var. sachlainense, 2n=42; E: G. pentapetalum, 2n=14. Arrows indicate satellite chromosomes. Bar represents 5ƒÊm. chromosomes were divided into three groups: 13 metacentric pairs, six submetacentric pairs, and two subtelocentric pairs (Figs. 2C, 3C). Satellites were found on the short arms of the two submetacentric pairs. The karyotype was formulated as 2n=42=26m+8sm- 4tsm+4st. (4) Geum macrophyllum Willd. var. sachalinense (Koidz.) Hara Three plants from the botanic garden of Hokkaido University were studied (Table 1). All had 2n=42 (Fig. 1D), which was consistent with the previous reports (Gajewski 1948, 1949, 1957, Taylor and Brockman 1966). Metaphase chromosomes in the one plant studied ranged 1993 Karyotypes on Five Species of Japanese Geum 317 Table 4. Measurements of somatic metaphase chromosomes of Geum japonicum t: satellite. Table 5. Measurements of somatic metaphase chromosomes of Geum macrophyllum var. sachalinense t: satellite. 318 Yoshikane Iwatsubo and Naohiro Naruhashi Cytologia 58 from 0.9ƒÊm-2.2ƒÊm in length and 1.0 to 3.5 in arm ratio (Table 5). These were divided into three groups: 13 metacentric pairs, six submetacentric pairs, and two subtelocentric pairs (Figs. 2D, 3D). Satellites were found on the short arms of the two submetacentric pairs. The karyotype was expressed as 2n=42=26m+8sm+4tsm+4st. Fig. 3. Idiograms of five species of Japanese Geum. A: G. aleppicum (6x), B: G. calthaefolium var. nipponicum (6x), C: G. japonicum (6x), D: G. macrophyllum var. sachalinense (6x), E: G. pentapetalum (2x). Table 6. Measurements of somatic metaphase chromosomes of Geum pentapetalum t: satellite. Table 7. Karyotypic data of five Japanese Geum species 1993 Karyotypes on Five Species of Japanese Geum 319 (5) Geum pentapetalum (Linn.) Makino Four plants were examined (Table 1) . All had 2n=14 (Fig. 1E), which was consistent with the previous reports, under the name of Sieversia pentapetala (Linn .) Greene, of Sokolovs kaya (1960, 1963). Karyotypes of the four plants showed no marked differencies . Chromo somes at somatic metaphase ranged from 0 .8ƒÊm-1.5ƒÊm in length, and 1.1 to 8.8 in arm ratio (Table 6). The somatic chromosome complement was classified into three groups: four me tacentric pairs, two submetacentric pairs , and one telocentric pair (Figs. 2E, 3E). The telocentric pair had a large satellite on the short arm . The karyotype was thus formulated as 2n=14=8m+4sm+2tt. Discussion Chromosome numbers of the Japanese Geum species examined were 2n=14 for G. pen tapetalum, and 2n=42 for G. aleppicum, G. calthaefolium var. nipponicum, G. japonicum and G. macrophyllum var. sachalinense, all of which are in accordance with the earlier reports (Fedorov 1969). Since the basic chromosome number of this genus has been recognized as x=7 (Darlington and Wylie 1955), the chromosome counts of 2n=14 and 2n=42 in the Jap anese Geum are diploid and hexaploid levels, respectively. This study is the first report of karyotype analysis in this genus (Table 7). Although these Geum species had gradual and symmetric karyotypes in common, the following similarities or differencies in karyotypes are as follows: (1) Geum calthaefolium var. nipponicum has one pair of subtelocentric satellite chromosomes, whereas the other three hexaploid species have two pairs of submetacentric satellite chromosomes. Thus, these hexaploid species fall into two groups based on the num ber of their satellite chromosomes or the centromeric position. (2) The three species, G. alep picum, G. japonicum and G. macrophyllum var. sachalinense, have remarkably similar karyo types, uniformly formulated as 2n=42=34(m+sm)+4tsm+4st. (3) The satellite chromo somes of G. pentapetalum are by far the most asymmetric, and differ from the other Japanese Geum in this regard. This comparative study of karyotypes among the Japanese Geum species reveals that G. aleppicum, G. japonicum and G. macrophyllum var. sachalinense are closely related, whereas G. calthaefolium var. nipponicum is distinct among the hexaploid Geum. Also, the diploid G. pentapetalum is quite distinct from the other Japanese Geum species studied. Sometimes, G. calthaefolium var. nipponicum has been placed in Acomastylis (e.g. Bolle 1933, Hara 1936), while G. pentapetalum has been treated as Sieversia (e. g. Bolle 1933, Okuyama 1977, Shimizu 1982, Sugimoto 1978). Our karyotypic data agree with these taxonomic treatments. Summary Karyotypic studies were done on five Japanese species of Geum. These Geum species had gradual and symmetric karyotypes in common. Karyotypic formulae of the five species were the following: G. aleppicum (6x), 2n=42=24m+10sm+4tsm+4st; G. calthaefolium var. nipponicum (6x), 2n=42=32m+6sm+2st+2tst; G. japonicum (6x), 2n=42=26m+8sm+ 4tsm+4st; G. macrophyllum var. sachalinense (6x), 2n=42=26m+8sm+4tsm+4st; G. pen tapetalum (2x), 2n=14=8m+4sm+2tt. The study of karyotypes of the Japanese Geum species reveals that G. aleppicum, G. japonicum and G. macrophyllum var. sachalinense have close re lationship, whereas G. calithaefolium var. nipponicum is distinct from the other hexaploid Geum. Also, G. pentapetalum was quite distinct from the other Japanese Geum species studied. 320 Yoshikane Iwatsubo and Naohiro Naruhashi Cytologia 58 Acknowledgment The authors are grateful to Dr. Hideki Takahashi for providing G.