HETEROSIS IN RED CLOVER Rolf Manner Agricultural Research Centre, Department of Plant Breeding, Jokioinen Received March 3, 1963 Red clover ( Trifolium pratense L.) is one of the most important fodder crops in northern Europe. The importance of red clover in Finland is emphasized by Paatela (10) and Valle (25). Extensive plant breeding work is carried out with red clover in northern Europe, i.e. in Denmark, Finland, Norway and Sweden (4). The charac- teristics of the red clover material grown in Finland are investigated by Paatela (11) The plant breeding methods used in Finland for red clover are inter alia de- scribed by Pohjakallio (13, 14, 15), Pohjakallio, Multamäki & Nuorvala (16) and Paatela (12). Red clover is on the whole a cross-fertilizing plant, but several investigators have also found self-fertility in red clover. The self-fertility varies very much and in many cases it seems to be pseudofertility (2, 5,6, 17, 19, 20, 23, 26, 27, 28). A more or less pronounced heterosis in red clover was noted in certain characters by Williams (29), Wexelsen (26), Salamov (18) and Lielmanis (7). Bulk crossing of red clover for exploitation heterosis has been tried by Salamov (18), Lielmanis (7) and Paatela (verbal information). Material and methods The present author found it of practical interest for plant breeding and plant husbandry to undertake experiments to explore the practical possibilities of making bulk crosses and mixtures between red clover strains, and of investigating the possi- bilities of obtaining practically usable strains of crosses by a breeding method for combining ability. The present investigation has been carried out at the Plant Breeding Institute Gulläker, Hammenhög, Sweden, in connection with the practical plant breeding work at the station. »Strain» = marketed variety, local strain or number variety. https://www.c-info.fi/en/info/?token=19GbHTz0fxqYYz93.ea0o1LN-cnU_T02xCbdmTA.8tmEpyYn694xLliaheLgnmV2Uh87r8bs9xCvJ0cj7Se0X0brlh95-qRd9FKmipCZ-7Ppa70_bzVZi51gPcaDRCcRastGEV4k5Kbg2jBfBNIzZghq5XpaTWS7OE18PbSnbvaSpKPEwxVWnTkassqgWCBaVBG3K33iFg 48 The following marketed strains are used in the present investigation as parents; the Swedish strains: Merkur, Monark, Essi, Lindby 11, Resistenta, Silo, and Bara and the Danish strains Hinderupsgaard, Hjelm, C tofte, and Daeno 111. In addition a number of experimental strains in breeding work were also used in crosses. As space does not allow the describing of all the strains used in crosses, the author wishes to state that the material is representative of a broad Swedish and in some part Danish material in red clover. In addition, the author will endeavour to show that this is an average material of diploid red clover cultivated in the south of Sweden. The number of trials (comparisons) and crosses (combinations) is given in the tables. In the tables the yields are given for different harvest years, »n» = number of harvest seasons. Generally speaking it may be said that the number of crosses is rather large, but that the different crosses have undergone trials for only a short time. The latter is due to the fact that the yield experiments in red clover require much time and that unfortunately the time period has been limited. The trials were carried out at the experimental farm Gulläker in the years 1951—58. The plots have in all cases been 10 m 2 and the number of replications have in most cases been 3—5. The largest difference in flowering time has been 12 days between strains tried in the same experiment (Essi-Merkur) (8), in all other com- parisons the differences have been smaller. The question hasrisen whether the differ- ences in earliness have influenced the possibilities of comparing the earliest and latest developing strains with each other. The above mentioned strains have, how- ever, been compared in a number experimental series (1, 3, 21, 22). In the govern- mental experimental programme SJF 210 the above strains are also compared in the same experiments. Such an experimental series has also been carried out at Gulläker in the years 1954—58 (21). These trials have taken place in the same experimental fields as the experiments of the present study. The harvest has in the present study been taken in the same time as in the comparable governmental trials at Gulläker and the same methods have been used. The methods including the harvest time determination have consequently been officially approved. The harvest has been carried out when the first flowers have appeared in Merkur i.e. when the earlier strains have had a number of flowers. Reasons for the comparison of strains of different earliness in the same governmental experiments have been interalia, that the red clover strains differ greatly, i.e., the earliness of the different plants in the same strain varies (24). Further it has been expected that the errors caused by the differ- ence in earliness are smaller than the errors which would arise if the comparisons were undertaken in different experiments, or if different strains in the same experi- ment were harvested on different days. Further it is expected that the errors will be small or none if two harvests per season are taken. These arguments have also been accepted in the present investigation. All possible measures were taken to avoid influence from the border effect. All weights refer to fresh green matter weighed immediately after harvest with an accu- racy of 100 grams. The seeds used for the experiments were received, in the case of the marketed strains, from the breeders. The crosses were produced by growing the strains in alternating rows or alternating plots. The seed was then harvested separately for 49 the different crosses. The number of replications varied from 3 to about 20 in the seed cultivations, being in most cases 3—5. As the parents had good normal fertility crosses between the strains grown together as well as within the strains occurred. The »hybrid seed» production was consequently not based on male sterile material. Two different types of »hybrid seed» were produced, namely: 1. Separate bulk crosses i.e. two or more strains are crossed together, but only the seed for the mother plots is included. For example, if the strains A and B'are crossed together the seed harvested is a mixture of about 50 % A x B and 50 % A X A, and if three strains A, B and C are crossed together the harvested seed will be a mixture of about 33.3 % A x B, 33.3 % A x C and 33.3 % Ax A. 2. Reciprocal bulk crosses i.e. two or more strains are sown together either in different rows or the seed is mixed before sowing the seed cultivation. According to the above mentioned examples the seed harvested will be about 50 % A x B, 25 % A x A and 25 % B X B, if two strains are crossed together. If three strains are crossed together, the seed will be about 22.2 % A x B, 22.2 % A x C, 22.2 % B x C, 11.1 % A x A, 11.1 % B x B and 11.1 % C X C. Of the two forms of bulk crosses three different types of crosses have been investigated, namely, with 1. two strains involved, 2. three strains involved and 3. more than three strains involved. The percentage of crosses obtained is discussed in an article on beets (9), In cases where two strains are grown in alternating rows the percentage of crosses will be d; 50, if no better fertility is to be found between different strains than within the strain and the earliness of the strains being about the same. In many cases in the present investigation selection for persistence in the gen- eration for seed cultivation was performed. This selection was a natural selection for persistence, performed in such a way that seed was taken in two or three year old seed cultivations established and treated in the same manner as earlier described, or plants were chosen from old plantations. The mixtures of seed were made in the laboratory in such a manner that equal quantities of germinative seeds were mixed and the seed was sown in seed cultiva- tions; the harvested seed from the seed cultivation was then used for green matter trials. To obtain the largest possible opportunities for comparison, all values in the calculation were performed as dr values in comparison with the check. The present paper does not give the names of the best crosses nor the names of the parents used in them, as these crosses may possibly be of commercial interest to my former em- ployer Otto J. Olson & Sons Ltd. Comparison between mother strains and progeny The first point considered in planning the present investigation was whether the bulk crosses have on an average better or worse yields than the parental strains? The parental strains are the best average strains grown in the area in which the experiments were conducted. 50 In the present material a total of 36 crosses have been tested. On an average the bulk crosses gave larger green matter yields in all the four harvests investigated. Since the different bulk crosses are not very much tested it has not been possible to draw conclusions as to the performance of individual crosses. Table 1. Comparisons between mother strains and progeny First harvest year Second harvest yearNuipber of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield All bulk crosses Mothers 69 44 Kilograms per hectare 28322 16016 44338 20451 13429 33880 Relative value 100 100 100 100 100 100 Progeny 36 69 44 Kilograms per hectare 28886 17998 46884 20747 13917 34664 Relative value 102 112 106 101 104 102 Table 2. Crossings in which two strains are crossed together Number First harvest year Second harvest year of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield All bulk crosses Parents 14 Kilograms per hectare 28947 Relative value 100 Progeny 8 14 Kilograms per hectare 30231 Relative value 104 Separate bulk crosses Parents 10 Kilograms per hectare 27351 Relative value 100 Mothers 10 Kilograms per hectare 28403 Relative value 100 Progeny 4 10 Kilograms per hectare 29939 Relative value (parental mean) 109 Relative value (mothers) 105 Reciprocal bulk crosses Parents 4 Kilograms per hectare 30439 Relative value 100 Progeny 4 4 Kilograms per hectare 30961 Relative value 102 14 17428 46375 22291 11622 33913 100 100 100 100 100 14 17677 47908 21996 12127 34122 101 103 99 104 101 10 17545 44896 22880 12748 35628 100 100 100 100 100 10 16069 44472 22005 13418 35423 100 100 100 100 100 10 17675 47614 23155 13622 36777 101 106 101 107 103 110 107 105 102 104 4 19636 50075 20816 11743 32559 100 100 100 100 100 4 17683 48644 19096 11185 30281 90 97 92 95 93 51 The reason for a larger number of comparisons in the first harvest year (Table 1) lies partly in the fact that a number of second year experiments were not harvested by the time the results were collected in to tables and partly in the fact that a number of second year experiments could not be harvested because the plants were killed by clover rot (Sclerotinia trifoliorum Eriks.). Dependence on number of strains crossed together In the present investigation an effort has been made to cross two, three or more strains together. The mean results are given in Table 2 (two strains crossed together), Table 3 (three strains crossed together) and Table 4 (several strains crossed together). The results show that no big differences are to be found between crosses in which 2 or 3 strains are crossed together if a comparison is made with the Table 3. Crossings in which three strains are crossed together Number First harvest year Second harvest year of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield Parents 12 9 Kilograms per hectare 29321 17772 47093 24538 13444 37982 Relative value 100 100 100 100 100 100 Mothers 12 9 Kilograms per hectare 29406 17987 47393 25182 13421 38603 Relative value 100 100 100 100 100 100 Progeny 4 12 9 Kilograms per hectare 30733 18749 49482 26103 13359 39462 Relative value (parental mean) 105 106 105 106 99 104 Relative value (mothers) 105 104 104 104 100 102 Table 4. Crossings in which many strains are crossed together First harvest year Second harvest yearNumber of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield Parents 8 6 Kilograms per hectare 27654 15427 43081 i 106-19 13462 33081 Relative value 100 100 100 100 100 100 Mothers 8 6 Kilograms per hectare 27420 14908 42328 19255 12828 32083 Relative value 100 100 100 100 100 100 Progeny 4 8 6 Kilograms per hectare 26483 15092 41575 14346 12560 26906 Relative value (parental mean) 96 98 97 73 93 81 Relative value (mothers) 97 101 98 75 98 84 52 parental mean, but if a comparison is made with the mothers, the bulk crosses, based on three strains, have shown results that are not as good as the bulk crosses based on only two strains. The result of this last mentioned comparison is further- more in agreement with the results obtained with bulk crosses in which many strains were crossed together (Table 4). This table shows that the yields of the bulk crosses based on many strains have been considerably smaller in comparison with those of the parents than those of bulk crosses, which are based on only a few strains. The yields in the second harvest year are much smaller relatively for the bulk crosses based on many strains than those based on only two or three strains. Comparisons between separate and reciprocal hulk crosses In Table 5 a comparison is given between separate and reciprocal bulk crosses. The number of combinations and trials was much larger for the separate bulk crosses. The separate bulk crosses have on an average given higher green matter yields than the means of the parents. The corresponding comparison for the reciprocal bulk crosses shows the average green matter yields for both harvest years to be smaller than for the parents. Table 5. Comparison between separate and reciprocal bulk crosses First harvest year Second harvest yearNumber of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield Separate bulk crosses Parents 62 39 Kilograms per hectare 28256 16034 44290 20592 13525 34117 Relative value 100 100 100 100 100 100 Progeny 36 62 39 Kilograms per hectare 28969 18252 47221 21473 13985 35458 Relative value 103 114 107 104 103 104 Reciprocal bulk crosses Parents 7 5 Kilograms per hectare 28906 15852 44757 19354 12682 32036 Relative value 100 100 100 100 100 100 Progeny 3 7 5 Kilograms per hectare 28151 15746 43897 15090 13385 28475 Relative value 97 99 98 78 106 89 The difference between the reciprocal and the separate bulk crosses is that the reciprocal bulk crosses are (Pj + P 2) + (Pl?=iP2), while the separate bulk crosses are P 1 -f (P x x P 2). The present author found at an early stage an indication that the separate bulk crosses would give the best practical results if the best strain the strain with the most desirable characteristics was used as mother strain. In this connection it must however be pointed out that the separate bulk crosses, not only 53 in comparison with the parental mean (Table 5) but also in comparison with the mothers, have given higher green matter yields (Table 2). The influence of selection on bulk crosses From the results discussed above it appears that the bulk crosses have on an average given larger green matter yields than both the mothers and, on an average, the parents, if compared in every individual case. It is further indicated by the data given previously that the average performance is not as good in the second harvest year as in the first. The present author consequently came at an early stage to be interested in trying a selection for persistence parallel with the production of the Table 6. Comparisons between parents and progeny at bulk crossing and selection Number First harvest year Second harvest year of combi- n First Second Total n First Second Total nations harvest harvest yield harvest harvest yield Parents 13 10 Kilograms per hectare 26958 14812 41770 18468 13607 32075 Relative value 100 100 100 100 100 100 Mothers 13 10 Kilograms per hectare 27246 15125 42371 18168 14190 32358 Relative value 100 100 100 100 100 100 Progeny 5 13 10 Kilograms per hectare 28349 18386 46735 22350 14869 37219 Relative value (parental mean) 105 124 112 121 109 116 Relative value (mothers) 104 122 110 123 105 115 seed of bulk crosses. The selection was carried out either in plantations or in sown stands. In the latter case three-year old sown stands were harvested for seed after natural selection. In planted stands selection was performed in the second or third year. In Table 6 are given the average results of separate bulk crosses performed with selected plants of different strains. Summary The possibilities of utilizing heterosis in reciprocal and separate bulk crosses and through selection parallel with bulk crossing in red clover are discussed. The reciprocal bulk crosses have not reached the average green matter performance of the pure strains in the crosses. The performance in the second harvest year was on an average not as good as in the first harvest year. On an average, the separate bulk crosses have given higher green matter yields than both the mothers and the parental 54 means. The yield performance was better relatively in the first harvest year than in the second harvest year in comparison with the mothers and with the parental means. If the bulk crosses are performed on plants selected for persistence the green matter yields have on an average been relatively higher than in other bulk crosses. The yields in the second harvest year have been comparatively better than in the first harvest year. Acknowledgements. The present investigation has been carried out at the Gull- äker Plant Breeding Institute, Hammenhög, Sweden, in the years 1949—58. My best thanks are due to my employer during the years 1949—58, Otto J. Olson & Sons AB and their late Managing Director Mr. Gottfrid Olson, B. Agr. My most sincere thanks are due to Professor Onni Pohjakallio, Dr. Agr. and Forestry, who has read the manuscript, to Mr. Erik Olsson and Miss Brigitte Bretschneider, who have helped me with the carrying out of the experiments, to my wife Mrs. Anita Manner, who has helped me with the calculations and treatment of the numerical material, and to Mr. C. Montagu Evans, M. A. (Cantab.) and Mr. Kalevi Multamäki, Dr. Agr. and Forestrg, for the linguistic revision. LITERATURE (1) Bengtsson, A. & Larsson, N. G. 1958, Stamförsök med slättervallväxter i södra Sverige under ären 1948 1957. K. lantbr. högsk. och stat. lantbr.förs. Stat. jordbr.förs. 112: I—2B. (2) Fergus, E. N. 1922. Self-fertility in red clover. (A report of the progress in the attempt to secure self-fertile lines in red clover). Kentucky Agr. Exp. St. Circ. 29: 19 36. (3) Jul£n, G. 1951. Svalöfs Silo rödklöver en ny klövertyp. Sveriges utsädesför, tidskr. 61: 347 355. (4) Jul£n, G. & Äkerberg, E, 1951. Vallväxter. Svensk växtförädling 1: 423 506. Natur och Kultur, Stockholm. (5) Kirk, L. E. 1925: Artifical self-pollination in red clover. Sci. Agr. 5: 179—189. (6) — »— 1933. The progeny test and methods of breeding appropriate to certain species of crop plants. Amer. Nat. 67: 515 531. (7) Lielmanis, J. 1955. Clovers of the Latvian SSR. (Russ,). Zemledelie 1955: 52 55. (8) Manner, R. 1952. Nägra erfarenheter rörande äterväxtförmägan hos vära vanligaste vallväxtarter och -stammar. Medd. Gulläkers växtförädlingsanst. 9—10: 219—224. (9) — »— 1960. Studies in bulk crosses between some beet strains. J. Sci. Agric. Soc. Finl. 32: 199 210. (10) Paatela, J. 1953. Maamme heinänurmien botaanisesta koostumuksesta. Suomen maatal. tiet. seur. julk. 79(3): 1 128. (11) —*— 1962. Characteristics of some diploid and tetraploid varieties of the late red clover Trifolium pratense v. subnudum subv. serotinum. Ibid. 99(4): 1 31. (12) —*— 1962. Viljelyskasvien alkuperä ja jalostaminen. Maanviljelysoppi 2: 1 33. Porvoo Helsinki. (13) Pohjakallio, O. 1938. Beiträge zu den ziichtungstechnischen Untersuchungen bei der Veredlung des Rotklees. J, Sei. Agric. Soc. Finl. 10:67 77, (14) —*— 1939. Kasvinjalostusbiologisia tutkimuksia apilamädästä. S. tiedeak. esit. ja pöytäk. 1939: 115 128. Ylipainos. (15) —»— 1939. Untersuchungen iiber den Kleekrebs und seinen Anteil am Verschwinden des Klees in Kleegrasgemischen. Pflanzenbau 16: 136 160, 201—205. (16) Pohjakallio, O. & Multamäki, K. & Nuorvala, S. 1937. Puna-apilan jalostusteknillisiä tutki- muksia. Referat: Veredlung des Rotklees. Ziichtungstechnische Untersuchungen. Valt. maatal. koetoim. julk. 13: 1 66. 55 (17) Renard & Lappo, A. 1928. Beiträge zur Kenntnis der Biologic des Blfihens von Rotklee (Trifolium pratense L.) verschiedener Herkunft. (Russ, with summary in English). Ann. der Weiss- ruthen. Staatl. Akad. f. Landw. in Gorky. 6: 201 219, (18) Salamov, A. 1953. Breeding research at the North Osetian State Breeding Station (Russ.). Selektsija i semenovodstvo 1953: 19 24. (19) Schieblich, J. 1939: Selbstfertilität bei Rotklee (Trifolium pratense L.). Ztichter 11: 89 90. (20) Schlecht, F. 1922. Untersuchungen fiber die Befruchtungsverhältnisse bei Rotklee (Trifolium pratense). Z. f. Pfl. zficht. 8: 121 157. (21) Statens Jordbruksförsök, 1954 59. SJF 210. Försöksplaner 1954—59. (Mimeographed). (22) Statens Jordbruksförsök 1956. Planer för Hushällningssällskapens lokala växtodlingsförsök är 1957. 1 57. Stockholm. (23) Sylven, N. 1929. Om rödklövems självfertilitet. Nord. jordbr. forsk, foren. 1929: 697 712. (24) Umaerus, M. & Äkerberg, E. 1959. Till frägan om typ- och stambeteckning hos rödklöver. Sv. utsädesf. tidskr. 69: 111 126 (With English summary). (25) Valle, O. 1957. The problem of red clover seed production in Finland. J. Sci. Agric. Soc. Finl. 29: 177-184. (26) Wexelsen, H. 1945. Studies on fertility, inbreedingand heterosis in red clover (Trifolium pratense L.). Skrift. Norske Vidensk.-Akad. Mat.Naturv. Kl. Oslo 1: 1 141. (27) Williams, R. D. 1925. Studies concerning the pollination, and breeding of red clover. Welsh Plant Breecf. Sta. Bull. Ser. H. 4: 1 58. (28) —»— 1931. Self- and cross-sterility in red clover. Welsh Plant Breed. Sta. Bull. 12: 181 208. (29) —*— 1937. Heterosis in red clover. Effect of in-breeding in F 2 and F, populations. Welsh J. Agric. 13: 172-190. SELOSTUS; HETEROOSI PUNA-APILASSA Rolf Manner Maatalouden tutkimuskeskus, Kasvinjalostuslaitos, Jokioinen Vuosina 1950—56 on Gulläkerin kasvinjalostuslaitoksella suoritettu puna-apilakantojen välisiä risteytyksiä. Täten saatuja risteytymiä on kokeiltu kantakokeissa mainitun kasvinjalostuslaitoksen koe- kentillä vuosina 1951 58. Näitä risteytymiä on verrattu vanhempiinsa. Yhteensä on kokeiltu 36 eri risteytysyhdistelmää. Risteytymät ovat keskimäärin antaneet suurempia tuorerehusatoja kuin van- hempansa. Parhaimmiksi näyttävät tuorerehusadot muodostuneen silloin, kun risteytymien siemen on otettu valioyksilöistä tai vanhasta nurmesta. Viimeksi mainitussa tapauksessa apilan kestävyyskin näyttää parantuneen huomattavasti. Se on nimittäin tällöin pystynyt antamaan suhteellisesti suu- rempia tuorerehusatoja toisena kuin ensimmäisenä satovuotena.