2016 Volume 91 Issue 1 Pages 41-46
Sakamura (1918) reported the discovery of a polyploid series among eight species of the genus Triticum; this series consisted of 2x, 4x and 6x species with 2n = 14, 28 and 42 chromosomes, respectively. He mentioned in this article that all the materials he used were gifted by T. Minami of the same department of Hokkaido University, Japan. In addition to carrying out an extensive collection of cereal germplasms in the period 1914 to 1916, Minami wrote on October 7, 1915 to K. A. Flaksberger, a wheat taxonomist at the Bureau of Applied Botany, Saint Petersburg, Russia, requesting seeds of Russian wheat and other cereals. He sent Flaksberger a letter of acknowledgement for seed stocks on May 19, 1916; thus, the requested seed package must have arrived from Flaksberger at some time between October 7, 1915 and May 19, 1916. Based on the available documents, there was a considerable period of time between these seed stocks reaching Minami and Sakamura’s publication of the chromosome numbers with the discovery of polyploidy. In fact, the wheat species identified by Flaksberger (1915) and those studied by Sakamura (1918) were identical except for two wild species which appeared only in Flaksberger’s list. The available information supports a proposal that the wheat species used by Sakamura (1918) in his discovery of polyploidy, and later by Kihara (1924, 1930) in his genome analysis, originated from Flaksberger’s collection.
According to the information provided by the Hokkaido University Archives, Sakamura was born in Hiroshima, Japan in 1888. He graduated from the College of Agriculture, Tohoku Imperial University (now the Faculty of Agriculture, Hokkaido University) in 1913 and then enrolled in its Graduate School. He was appointed Assistant Professor of the College of Agriculture, Hokkaido University in November, 1918, and received an Overseas Scholarship from the Ministry of Education, National Government to study in the USA and Europe from December, 1918 to March, 1921. He was promoted in December of that year to Professor in charge of the Second Laboratory of Botany in the College of Agriculture, Hokkaido University, and moved in April, 1930 to the Department of Botany in the newly established College of Science in the same university.
Sakamura’s groundbreaking study to determine the correct chromosome numbers in wheat species leading to the discovery of polyploidyIn July, 1918, when Sakamura was a special scholarship graduate student at the College of Agriculture, he reported the correct chromosome numbers of wheat species, and discovered polyploidy in the genus Triticum (Sakamura, 1918); this report provided the basis for modern wheat genetics. This epoch-making study was published as a four-page article in the Botanical Magazine, Tokyo. In two years, we will celebrate the centennial of his groundbreaking work. Here, I wish to report information on the origin of Sakamura’s wheat stocks, which were subsequently used by Dr. Hitoshi Kihara for his well-known genome analyses of wheat (Kihara, 1924, 1930).
Sakamura published his 1918 article in German under the title “Kurze Mitteilung über die Chromosomenzahlen und die Verwandtschaftsverhältnisse der Triticum-Arten”, that is, “Short report on the chromosome number and phylogenetic relationship of Triticum species” in English translation. In this article, he presented the results of his chromosome observations in eight wheat species (Table 1).
| Species | Haploida) | Diploid | Species | Haploida) | Diploid |
|---|---|---|---|---|---|
| T. vulgare | 21 | 42 | T. durum | – | 28 |
| T. compactum | – | 42 | T. polonicum | – | 28 |
| T. Spelta | – | 42 | T. dicoccum | – | 28 |
| T. turgidum | – | 28 | T. monococcum | – | 14 |
He determined chromosome numbers mainly from mitotic divisions in root tip cells, as he found more active cell division in the root tip than in young spikes. Meiotic chromosome number (n) was analyzed in only one species, T. vulgare (now called T. aestivum), whereas he determined the somatic 2n chromosome number in all eight species. The analyses showed that one species had 2n = 14, four species had 2n = 28 (i.e., double the number of the first species), and the remaining three species had 2n = 42 (i.e., triple the number of the first species). These results provided the first indication of polyploidy in the wheat genus Triticum.
In the 1918 article, Sakamura acknowledged Prof. Takajiro Minami of the First Laboratory, Department of Agronomy for his kindness in providing the wheat stocks used in his study. These stocks were derived from the germplasm collection made by Minami, the director of the University Farm of Hokkaido University at that time; later, Minami served as the First Dean of the College of Agriculture and then as the Second President of Hokkaido University.
T. Minami’s germplasm collection of cerealsFrom 1914 to 1916, Minami devoted himself to the collection of cereal germplasms, including wheat, barley, oats and corn. His activities have been traced by referring to carbon copies of letters sent to foreign institutions requesting crop seeds and from letters of acknowledgement following receipt of the seeds. These records had been stored in a corrugated cardboard box since that time in the Research Group of Botany and Agronomy, which is now chaired by Prof. Kazuto Iwama.
The documents reveal that Minami wrote to 63 institutions in five countries and regions, namely, Canada, Madagascar, Russia, Sri Lanka and the USA, requesting cereal seed samples and also seeds of other plant species. He received seed samples from 42 institutions: 98 wheat seed samples, 12 barley samples, 175 oat samples and 12 maize samples, along with 106 samples of other crops. Of the wheat samples, 78 were from 28 agricultural experimental stations or university farms in the USA and two were from the Canadian Central Experimental Farm; all the samples except one were recorded by their cultivar names. From this information on cultivar names, it is evident that the samples used by Sakamura were not derived from this part of Minami’s wheat collection. Minami also received 18 samples of wheat from Dr. K. A. Flaksberger of the Bureau of Applied Botany, Russia; however, no documents have been found that indicate the scientific or cultivar names of these samples. The possible relationship of the seed samples sent by Flaksberger to the wheat stocks used by Sakamura will be dealt with in the next section.
The same cardboard box also contained a notebook entitled “The Catalogue of Crop Specimens, 1915”. This notebook has a memo on its front page saying “The stocks of the First Laboratory, Agronomy Department, College of Agriculture, Tohoku Imperial University”. The notebook catalogues the germplasms for four cereals, wheat, rye, barley and oats; these are mostly listed by their scientific names, although some accessions have cultivar names. No indications of the origins of any stock are given. The number of accessions of each wheat species listed in the notebook is as follows: T. vulgare (= T. aestivum), 15; T. compactum, 2; T. spelta, 3; T. dicoccum, 4; T. durum, 7; T. polonicum, 5; and T. turgidum, 2. Thus, a total of seven species with 38 accessions are listed. It is not certain whether the wheat accessions studied by Sakamura are included in this collection, although seven of the eight species he studied are in the list; the missing species is T. monococcum.
Minami’s correspondence with FlaksbergerThe correspondence between Minami and Flaksberger was initiated by a letter dated October 23, 1913 from the latter to I. Hoshino, an Assistant Professor in Minami’s laboratory. In this letter, Flaksberger outlined a project at his institute for the collection of germplasm of foreign crops; he provided information on the main researchers engaged in this project and stated that as their collection did not include any Japanese crops they were requesting seeds of Japanese cultivars of wheat, barley and oats (especially wild type). This letter seems to have stimulated Minami to begin a collection of foreign germplasms of crops that were important for agriculture in Hokkaido. Minami started his germplasm collection in 1914 (as described in the above section), a year after the arrival of Flaksberger’s letter at Hokkaido University.
As director of the University Farm, Minami responded to Flaksberger’s request and immediately sent seeds of nine barley and three oat cultivars on December 27, 1913; these seeds had been harvested at the University Farm. In the same letter, he requested that Flaksberger also send the bulletin published by the Bureau of Applied Botany, Russia. On September 16, 1915, Minami sent Flaksberger additional seed samples of four wheat and six barley cultivars that he had obtained from other agricultural experimental station(s) on the main island of Japan. From May, 1914 to March, 1916, Minami received 14 issues of the Bulletin of the Bureau of Applied Botany from Flaksberger. In a letter of October 7, 1915, Minami asked Flaksberger to send him seeds of Russian wheat, barley, oats and sunflower, and received 18 wheat, 14 barley, three oat and one sunflower seed samples. Minami wrote a letter acknowledging receipt of these seed samples on May 19, 1916. Regrettably, we did not find any document that describes the cultivar or species names of these seed samples.
Possible time course of events between the arrival of Flaksberger’s seed samples in the laboratory of Minami and Sakamura’s publication reporting the chromosome numbers of wheat speciesAs described above, the seed samples sent by Flaksberger must have arrived between October 7, 1915 and May 19, 1916. The wheat accessions Sakamura obtained from Minami were all spring type; the practice in the Sapporo area was to sow seeds of this type of wheat in early May. The plants would have headed in early to mid-July and ripened in late August, according to the “Records of Spring Wheat Cultivation, Minami’s First Agronomy Laboratory, 1895”. If the seeds from Flaksberger arrived by early May, 1916, Minami could have propagated them during the 1916 crop season and harvested the seeds in September of the same year. Thus, it is possible that Sakamura received the Flaksberger-derived wheat stocks from Minami at the end of 1916 and germinated them in the laboratory in early spring, 1917. Root tips could have been collected from seedlings for cytological observation prior to transplantation into the field in early May, 1917. Chromosome analysis in root tip cells of all eight species could therefore have been completed between March and May, 1917. Analysis of meiotic chromosomes in pollen mother cells of T. aestivum, the only species with which this investigation was performed, could have been carried out in summer, 1917. This schedule provides sufficient time to obtain and analyze the results in 1917, and to submit an article to the Botanical Magazine, Tokyo for publication in July, 1918. Sakamura also made manual crosses between some tetra- and hexaploid species in summer, 1917 with the intention of investigating chromosome numbers in the offspring of hybrids. F1 seeds were obtained in the fall of 1917 and sown in April–May of 1918 by Sakamura, who was due to leave at the end of 1918 for an overseas study trip to the USA and Europe.
Hitoshi Kihara, who had enrolled on the graduate course in August, 1918, was given the F1 hybrids grown in the field by Sakamura. Kihara prepared slides with meiotic chromosomes from the pentaploid wheat hybrid, the first time this had ever been carried out, and was given a few minutes’ advice and guidance from Sakamura, who was just leaving for his overseas trip. Kihara later declared that this short period of guidance by Sakamura determined his subsequent career as a wheat geneticist (Kihara, 1973).
Identical wheat species studied by Flaksberger (1915) and Sakamura (1918)Dr. N. P. Goncharov, Head of the Wheat Genetics Laboratory, Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, attended the 12th International Wheat Genetics Symposium held in Yokohama, Japan in September, 2013. On this occasion, I asked him to examine records preserved in the Bureau of Applied Botany, Saint Petersburg, Russia (now named the N. I. Vavilov Institute of Plant Industry) regarding the wheat seed samples sent by Flaksberger to Japan between October 7, 1915 and May 19, 1916. Although I did not receive any information on this matter from Dr. Goncharov, he kindly sent me his article “Konstantin Andreevich Flaksberger”, published in the Russian journal Studies of the History of Biology (Vol. 5, No. 3, pp. 106–108, 2013), with his personal English translation. He also attached a copy of the front page of the January-February issue (Vol. 8, 1915) of the Bulletin of Applied Botany, Russia, to which Flaksberger had contributed an article entitled “Manual of wheats” (Flaksberger, 1915). Dr. Goncharov also copied a figure entitled “Scheme of wheats’ genesis according to the latest investigations” from the article. In this figure, Flaksberger drew phylogenetic relationships among wheat species; this schema is shown here in tabular form (Table 2).
| Conspecies | Tr. monococcum L. | Tr. eu-dicoccoides Flaksb. | Tr. speltoides Flaksb. |
|---|---|---|---|
| Wild forms | Tr. monococcum var. aegilopoides | Tr. dicoccum var. dicoccoides | (unknown) |
| Cultivated forms with brittle rachis and hulled grains | Tr. monococcum var. cereale | Tr. dicoccum var. sementivum | Tr. Spelta |
| Cultivated forms with stiff rachis and naked grains | (unknown) | Tr. durum | Tr. vulgare |
| Tr. polonicum | Tr. compactum | ||
| Tr. turgidum |
Disputable relationships are omitted from the table.
Flaksberger classified the wheat genus into three conspecies, Tr. monococcum L., Tr. eu-dicoccoides Flaksb. and Tr. speltoides Flaksb., all of which were subdivided into wild and cultivated forms, although the wild form was absent from the conspecies Tr. speltoides. The wheat species listed in Flaksberger’s classification and the wheat accessions studied by Sakamura (1918) are compared in Table 3.
| Flaksberger (1915) | Sakamura (1918) | ||
|---|---|---|---|
| Conspecies | Species | Ploidy | Species |
| Tr. monococcum | Tr. monococcum var. aegilopoides | Diploid | (not studied) |
| “ | Tr. monococcum var. cereale | “ | T. monococcum |
| Tr. eu-dicoccoides | Tr. dicoccum var. dicoccoides | Tetraploid | (not studied) |
| “ | Tr. dicoccum var. sementivum | “ | T. dicoccum |
| “ | Tr. durum | “ | T. durum |
| “ | Tr. polonicum | “ | T. polonicum |
| “ | Tr. turgidum | “ | T. turgidum |
| Tr. speltoides | Tr. Spelta | Hexaploid | T. Spelta |
| “ | Tr. vulgare | “ | T. vulgare |
| “ | Tr. compactum | “ | T. compactum |
Sakamura showed his materials only with their species names.
With the exception of the wild species Tr. monococcum var. aegilopoides and Tr. dicoccum var. dicoccoides, all of the species listed by Flaksberger were the same as those used by Sakamura (1918) for his analysis of chromosome numbers. In addition, both workers used the same way of writing “T(r). Spelta” for the spelt-type hexaploid species.
The possible time course from the dispatch of seed samples by Flaksberger to the publication of the correct wheat chromosome number by Sakamura along with the complete concurrence of their species lists strongly suggest that the wheat stocks used by Sakamura (1918) were derived from Flaksberger via Minami, who had received the seed stocks between October 7, 1915 and May 19, 1916.
The use of Sakamura’s wheat stocks by Kihara for his genome analysisSakamura made the following interspecific crosses between tetraploid and hexaploid species: T. durum (4x) x T. vulgare (6x); T. turgidum (4x) x T. compactum (6x); and T. polonicum (4x) x T. spelta (6x). He grew the F1 plants in the field in 1918 and gave these hybrids to Kihara, who enrolled on the graduate course of Hokkaido University in early August of that year (Kihara, 1919). Kihara (1924) analyzed meiotic chromosome behavior in these pentaploid hybrids, and also examined an additional pentaploid hybrid between T. polonicum x T. compactum and two triploid hybrids, T. dicoccum (4x) x T. monococcum (2x) and T. aegilopoides (now T. boeoticum) (2x) x T. dicoccum, that he had produced himself. He reported a 7II + 7I chromosome configuration in the triploids and a 14II + 7I configuration in the pentaploids in most meiocytes at meiotic metaphase I (‘II’ and ‘I’ indicate a bivalent and a univalent, respectively). Those results led Kihara to conclude that diploid and tetraploid wheats have one set of seven chromosomes in common and that the tetraploids have an additional and unique second chromosome set; he further suggested that tetraploid and hexaploid wheat had two chromosome sets in common, and that hexaploids contained a unique third set. From these findings, Kihara (1930) postulated that a set of seven chromosomes constituted the basic unit of inheritance, and gave this set the name ‘genome’, using the terminology of Winkler (1920). He proposed the genome formulae AA, AABB and AABBDD for diploid (einkorn), tetraploid (emmer) and hexaploid (common) wheat, respectively. Subsequently, Lilienfeld and Kihara (1934) studied a new tetraploid species, T. timopheevi, discovered by Zhukovsky in 1923 (after Jakubziner, 1958), and found that it had a different second genome from that of emmer wheat; they designated a new genome formula, AAGG, for this species. This marked the completion of genome analysis of the genus Triticum. Later, Kihara (1954, 1957) extended his genome analysis to another genus, Aegilops, the closest relative of Triticum, assigning a genome formula to each of 11 diploid, 10 tetraploid and three hexaploid species.
Kihara established a new Laboratory of Experimental Genetics in the Department of Agricultural Biology, Faculty of Agriculture, Kyoto University in 1927. He obtained the following eight wheat species from Hokkaido University: T. monococcum var. vulgare, T. dicoccum var. liguliforme, T. durum var. reichenbachii, T. turgidum var. nigrobarbatum, T. polonicum var. vestitum, T. spelta var. duhamelianum, T. vulgare var. erythrospermum and T. compactum var. humboldtii (Tanaka, 1983). These species are identical to those used by Sakamura (1918), suggesting that Flaksberger’s wheat stocks played an essential role in both the discovery of polyploidy by Sakamura (1918) and the genome analysis by Kihara (1924, 1930).
Social conditions in Russia when Flaksberger sent the wheat seed samples to MinamiRussia’s involvement in World War I from 1914 to 1917 triggered considerable social instability in the country, which resulted in the February and November Revolutions of 1917. As a consequence of these uprisings, the Russian Empire collapsed. Inevitably, this social instability also affected research activities at the Bureau of Applied Botany in Russia; this is evident from the number of research bulletins sent to Hokkaido University at Minami’s request: eight issues in 1914 (dated March, April, May, June, July, August and December, with one of unknown publication date), four in 1915 (January–March, April–May, June and August–September), two in 1916 (January and March) and none thereafter. It is something of a miracle that the seeds sent by Flaksberger between October, 1915 and May, 1916 were transported from Saint Petersburg to Sapporo along the Siberian Railway, a distance of more than 10,000 km, under such extremely difficult social circumstances to safely reach the hands of Minami and thus to have contributed to Sakamura’s and Kihara’s groundbreaking studies. It is worth noting that some stocks thought to be derived from Flaksberger are still actively used in Japan as testers for identifying the plasmon type (= cytoplasmic genome) of all species in the Triticum-Aegilops complex, contributing to plasmon analysis of this complex (for reviews, see Tsunewaki, 1996, 2009).
Concluding remarksIn conclusion, the wheat stocks that Minami obtained from Flaksberger between October, 1915 and May, 1916 were used in Sakamura’s seminal work that led him to discover polyploidy in wheat and in Kihara’s work on genome analysis. Watkins (1930) wrote in the introduction of his article “The wheat species: a critique” that wheat genetics entered a fresh phase with Sakamura’s discovery (1918) that the species might have 14, 28 or 42 chromosomes (diploid), and with Kihara’s work on the cytology of hybrids between species with different numbers of chromosomes. The seed stocks sent by Flaksberger from Saint Petersburg in 1915 or 1916 in response to a request by Minami at Hokkaido University opened the door of this new era. This story emphasizes the importance of both precise record-keeping with regard to the source of experimental stocks and maintenance of their living germplasm for future research.
PostscriptK. A. Flaksberger was arrested on June 28, 1941, about a year after N. I. Vavilov, and died on September 13, 1942 in Zlatoust prison hospital (after Goncharov, 2013).
Prof. E. Shikata, Member of the Japan Academy and Emeritus Professor of Hokkaido University, let me know about the documents kept in the laboratory of Prof. T. Minami cited here, and kindly arranged to send them to me through Prof. K. Iwama; I would like to express my sincere thanks to both of them for their generous help. This article is a modified translated version of my two articles published in the Annual Report of Hokkaido University Archives, No. 6 and No. 9 (Tsunewaki, 2011, 2014), with additional information about K. A. Flaksberger, which was supplied by Dr. N. P. Goncharov, Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences; I wish to acknowledge his kindness in supplying an English translation of his article (Goncharov, 2013) on Flaksberger’s life history. My thanks are due to the Hokkaido University Archives for their kindness in allowing me to publish this article. Useful advice and support have been received from Dr. M. Yamamoto and Dr. T. Inoue, Hokkaido University Archives, Dr. S. Ikeguchi, Hokuren Inc., and Dr. S. Nasuda, Faculty of Agriculture, Kyoto University, in different ways in preparation of this article.
