CYTOLOGICAL INVESTIGATIONS OF HYBRIDS AND HYBRID DERIVATIVES OF CREPIS CAPILLARIS AND CREPIS TECTORUM BY LILLIAN HOLLINGSHEAD University op California Publications in Agricultural Sciences Volume 6, No. 2, pp. 55-94, plates 1-3, 19 figures in text Issued January 21, 1930 University of California Press Berkeley, California Cambridge University Press London, England CYTOLOGICAL INVESTIGATIONS OF HYBRIDS AND HYBRID DERIVATIVES OF CREPIS CAPILLARIS AND CREPIS TECTORUM BY LILLIAN HOLLINGSHEAD INTRODUCTION Concurrently with the investigation of a lethal factor which manifested itself in F a hybrids of Crepis capillaris (L.) Wallr. and C. tectorum L. (Hollingshead, MS), a cytological investigation of the same material was undertaken. It included studies of the somatic and meiotic chromosomes of the parental species and of reciprocal hybrids, while the occurrence of several triploid hybrids (2n capillaris, n tectorum) provided material for a similar study of these and of some of their progeny. Meiotic phenomena in interspecific hybrids in this genus have been described by Collins and Mann (1923), M. Nawaschin (1927), and Babcock and J. Clausen (1929). The hybrids described in this paper resemble thase investigated by Babcock and Clausen, but differ from the comparable ones described by Collins and Mann and by Nawaschin in exhibiting a variable number of pairs in meiosis. Unpublished investigations on the number of bivalents and univalents in various other hybrids, by Dr. Margaret Maun Lesley, Miss Priscilla Avery, and the writer, have shown that this phenomenon is common in many interspecific hybrids in the genus. ACKNOWLEDGMENTS This study was undertaken at the suggestion of Professor E. B. Babcock, and the writer gratefully acknowledges her indebtedness to him and to Professor J. L. Collins for the material and for helpful advice and interest throughout the course of the work. 56 University of California Publications in Agricultural Sciences [Vol. 6 MATERIALS AND METHODS The eapillaris strain X which was used in most of the hybrids is of unknown origin, having descended from a stray plant picked up in the greenhoiise. The other strains were kindly supplied by Dr. J. L. Collins, and are of various origins. Two tectorum strains (1498, from Copenhagen Botanical Garden, and 1648, from Tomsk, Siberia) were involved in the hybrids examined. The 1498 strain appeared to be morphologically rather constant and had descended from one plant (Hollingshead, MS). Plants of the 1648 strain, from original wild seed, varied in leaf shape and time of maturity. Plate 1 shows typical eapillaris, tectorum, and F 1 hybrid plants at maturity. Sections of root tips fixed in chromacetic formalin as described by Hollingshead and Babcock (1929), and stained with Heidenhain's iron-haematoxylin were used for somatic chromosome studies. Meiotic phases were obtained from buds fixed in Carnoy's fluid, washed in absolute or 95 per cent alcohol, run through 80 per cent alcohol into 70 per cent, there to remain until examined. Material so fixed usually remained in good condition for at least a year. Considerable shrink- age was occasionally noted but on the whole the fixation was good. To make mounts from such material the florets were dissected out on a slide, crushed in a drop of aceto-carmine, the debris removed, a coverslip applied, and all excess fluid absorbed so that only a thin film remained and the coverslip held the loose pollen-mother-cells (PMC's) in place. The counts given later in the paper, each from one slide, will give some idea of the number of countable PMC's obtained from one bud in favorable material. In most cases the PMC 's remained in rows of a few to many cells and these were usually at the same stage of development. Pollen grains were mounted in acetocarmine and were counted "good" if the cytoplasm was well stained after twenty-four hours. All examinations were made with a Bausch and Lomb 90x apochromatic objective and Zeiss compensating oculars. The somatic chromosomes were drawn at a magnification of 3750, the meiotic at 2450, and the tetrads at 1250 and all were reduced one-third in reproduction. 1930] Eollingshead: Hybrids of Crepis capillaris and C. tectorum 57 SOMATIC CHROMOSOMES OF PARENTS AND HYBRIDS Crepis capillaris, as shown by Taylor (1925, 1926) and M. Nawa- schin (1925&), has six somatic chromosomes which are readily dis- tinguished as three pairs by their distinctive morphology. Nawaschin (1925&) has figured those of C. tectorum in which there are four pairs of chromosomes, each different. Somatic metaphase plates of these two species showing the same details of morphology as Taylor and Nawaschin have shown are given in figure A. For the sake of A Fig. A. Somatic metaphases of (1) C. capillaris, (2) C. tectorum. uniformity with Nawaschin 's system of designation, the three chromo- somes of capillaris will be designated A, C, and D, and those of tectorum A, B, C, and D (fig. A). The writer has no intention of signifying homology by these letters. Fig. B. Somatic metaphase of F, C. capillar is-tectorum. Nawaschin (1927) has figured the somatic chromosomes of the F x capillaris-tectorum hybrid and the writer's investigations confirm his observation that the tectorum D-chromosome in the hybrid is modified so that the satellite is visible no longer and the head is slightly enlarged (fig. B). The chromosomes of reciprocal hybrids are alike in this respect. 58 University of California Publications in Agricultural Sciences [Vol. 6 Measurements of chromosome length seem to indicate that this may not be the only chromosome change which accompanies hybridiza- tion. Measurements were made on all the chromosomes in each species and on capillaris chromosomes A and C and tectorum chromosomes A, B, and C, in reciprocal hybrids. Ten representatives of each chromosome from the protoderm or the row of cells immediately below it, lying quite or, in a few cases, almost horizontally, were drawn at the same magnification, projected by a lantern, and the figures so obtained were traced and the lengths measured by means of an opisometer run along the median line and back along a measured straight edge. Each chromosome was measured twice, or until two measurements agreed. The instrument, or the manipulation of it, gave measurements which varied rarely more than 0.2 cm. to a chromosome. All material from which measurements were made was fixed in the same fixative and treated in the same way. Table 1 gives the individual lengths and mean lengths obtained for chromosomes A and C of capillaris and A, B, and C of tectorum in the parental species and in reciprocal hybrids. The D-chromosomes of each species were not measured in the hybrids and so are not included in the table but the measurements on the species confirmed the opinion derived from observation that the D-chromosome of capillaris is slightly longer than that of tectorum.. Three different strains and a number of different plants were involved in the three sets of measurements of capillaris chromosomes and it might be that if there is any constant difference in the length of any of the capillaris chromosomes between the hybrids and the parental species, it might be the residt of this condition. All the tectorum measurements were from one strain but involved several plants. The data show that considerable variation occurs in the lengths of the same chromosome in different cells and that the chromo- somes from the protoderm (bracketed together at the top of each set of measurements) are on the whole somewhat longer than those in the cells beneath. There are in each set of measurements repre- sentatives from each of the two rows of cells from which chromosomes were measured. Table 2 shows the differences in mean lengths obtained for the various chromosomes between the parental species and reciprocal hybrids, the probable errors of the differences, and the relation of the differences to their probable errors. Most of the chromosomes showed no significant difference in mean length in parental species and in hybrids. The A-chromosome of 1930] Hollingshead: Hybrids of Crepis capillaris and C. tectorum 59 TABLE 1 Lengths of Ten Kepresentatives of Five Different Chromosomes of C. capillaris and C. tectorum from the Boot Tips of the Parental Species and Beciprocal Hybrids capillaris tectorum A C A B C Parental species 18.0 18.0 18.3 21.0 20.2 17.2' 16.6 16.8 18.0 17.5, > 8.5' 8.7 8.8 11.0 9.7 9.3 9.2 8.5 8.9 8.9, ► 19.6' 20.7 19.3 19.4 19.9^ 18.7 17.3 17.0 18.5 19.3, • 15. f 17.6 14.3 17.8 18.6 17.4 16.1 15.5 16.8 15.1, > 14.8' 15.1 15.5 14.1 14.0 13.2 14.2' 14.8 14.2 13.9, - Mean 18.16 9.15 18.97 16.43 14.38 tectorum 9 X capillaris tf 18.2' 20.1 17.0 16.3 17.8 16.0' 19.2 17.5 17.6 19.6, 10.8] 9.7 11.0 10.1 10.4' 8.9 9.8 10.5 10.7 9.3, » > 17.4 17.3 21.1 19.9^ 18.8 17.8 19.0 18.2 15.8 18.8, y 18.0' 17.0 16.4 18.0 17.5 19.3 17.1 16.7 16.8 16.2, • 13.5 1 12.8 13.2 13.1 13.4 14.2 12.7 13.3 11.5 11.8, Mean 17.92 10.12 18.41 17.3 12.95 capillaris 9 X tectorumd 1 19.0 18.5 19.2 20.1 17.3 16.0 20.3 16.9' 18.9 17.0, > ► 9.3 10.2 10.5 9.7 10.0, 8.8' 9.2 9.7 9.9 10.5, > > 18.3 17.7 17.2 17.5 18.4 18.9 19.5 16.3 15.3 16.7, > 18.81 16.4 16.1 15.8 15.7 16.0 16.8 16.6 16.6^ 15.4J > > 14.2' 13.8 13.7 11.9 13.7 11.8 13.1 13.4' 11.8 12.7, Mean 18.32 9.78 17.58 16.42 13.01 60 University of California Publications in Agricultural Sciences [Vol. 6 TABLE 2 Differences in Mean Lengths of Five Chromosomes in the Parental Species and Reciprocal Hybrids Chromosome Mean length in parents Hybrid Mean length in hybrid Parental —hybrid mean Probable error of difference Diff. Ed capillaris A 18.16±0.286J capillaris 9 tectorum 9 17.92±0.272 18.32±0.293 +0.24 +0.16 ±0.395 ±0.409 0.6 0.4 capillaris C 9.15±0.710| capillaris 9 tectorum 9 978±0.530 10.12±0.138 +0.63 +0.97 ±0. 189 ±0.205 3.3 4.7 lector urn A 18.97±0.229J tectorum 9 capillaris 9 18.41±0.298 17.58±0.255 -0.56 -1.39 ±0.376 ±0.344 1.5 4.0 tectorum B 16.43±0.286J tectorum 9 capillaris 9 17.30±0.188 16.42±0.191 +0.87 -0.01 ±0.343 ±0.354 2.5 0.03 tectorum C 14.38±0.135J tectorum 9 capillaris 9 12.95±0.162 13.01±0.183 -1.43 -1.37 ±0.211 ±0.228 6.7 6.0 tectorum had a shorter mean length in both hybrids, 1.5 and 4 times the probable error respectively, which in view of the small number of measurements cannot be said to be very significant. An increase in length in the C-chromosome of capillaris, 4.7 and 3.3 times the prob- able error in reciprocal hybrids, may be significant and might be attributed to hybridization if the same strain had been used in all measurements. The data on tectorum C showing a difference of over 6 times the probable error in each case would seem to indicate clearly that this chromosome was constantly shorter in the hybrids examined than in the plants of the species examined. The data can be considered only as of a preliminary nature, but they suggest that certain chromosomes in a hybrid may be constantly modified in size as well as in form. Where the difference is at all great the modification is in the same direction in reciprocal hybrids and would suggest that the change was a result of a mutual reaction between the chromosomes of the two hybrid complexes. Yet it is evident that all the chromosomes of a complex were not affected in the same way. It would appear worth while to investigate this and other suitable material more extensively in this connection. 1930] B oiling she ad: Hybrids of Crepis capillaris and C. tectorum 61 MEIOSIS IN THE PARENTAL SPECIES Stages prior to diaphase (terminology, Belling, 1928) have not been studied in either parents or hybrids. With the exceptions to be noted, meiosis proceeds similarly in the two parental species and the descriptions apply equally to both. Middle diaphase figures usually show chromosome pairs elongated and more or less loosely associated, as depicted for capillaris in figure C, 1. One of the pairs seems often to lie with an end on the nucleolus and in a number of cases in capillaris this pair has been determined as the middle-sized one. Since it is this pair which bears the satellites in somatic cells, this agrees / 2 Tig. C. (1) Middle and (2) late diaphase of C. capillaris. with the findings of those who have reported satellites associated with the nucleolus (cf . Kuhn, 1928) . Late diaphase, a stage evidently passed through rather quickly, for only occasionally does a slide show many such figures, is particularly favorable for showing size differences in chromosome pairs (capillaris, fig. C, 2). The shapes of the bivalent chromosomes are not constant. First metaphase (I-M) plates of the two species with three and four bivalents, respectively, are given in figure D, 1 and 2. Again the shapes of the bivalents are not constant. In capillaris it is usually possible to distinguish at least the smallest pair by its size, but this stage is less favorable than the one just described for determining size differences. All the first late anaphase (I-A) chromosomes show a longitudinal split similar to that figured later for hybrids. At second metaphase (II-M) and anaphase (II-A) the chromosomes resemble those in somatic divisions (fig. E, 1 and 2) and here they can often be distinguished from one another. With the exception of an occasional case of non-conjunction (3 out of 47 PMC's in one slide showed this condition), meiosis in the 62 University of California Publications in Agricultural Sciences [Vol. 6 tectorum plants examined proceeded normally and regular tetrads were formed. So much cannot be said for capiUaris, for in the X- strain of this species, which was the one most studied, meiotic irregu- larities occurred frequently. Three plants of this strain on which I-M counts were made showed higher percentages of irregularities in meiosis and tetrads than did any of the tectorum plants examined, but very noticeable differences in the amount of irregularities were evident between the various plants. The most frequent form of 1 2 Fig. D. Side views of meiotic metaphases of (1) C. capiUaris, (2) C. tectorum. 1 2 Fig. E. Second meiotic metaphase and anaphase of C. tectorum. irregularity was non-conjunction of the members of one pair of chromosomes but occasionally four chromosomes were unpaired. Non- conjunction occurred at diaphase (fig. F, 1 and 2) as well as at meta- phase (fig. G, 1 and 2). When one pair of chromosomes failed to conjugate, it was apparently not always the same pair (fig. F, 1 and 2). Abnormal elongation of bivalents and fragmentation at I-M occurred frequently in some slides (fig. H, 1 and 2). Table 3 shows the amount of non-conjunction observed in several representative plants of this species, each count having been made from a single bud. Though admittedly inadequate, the results may indicate that strains other than X are more nearly regular in meiotic phenomena, and this is upheld by pollen counts (below). Plants 1930] H oiling shead : Hybrids of Crepis capillaris and C. tectorum 63 Fig. F. Late diaphases of C. capillaris showing non-conjunction in different chromosome pairs. Fig. G. Meiotic metaphases of C. capillaris showing non-conjunction of (1) one-chromosome and (2) two-chromosome pairs. Fig. H. First meiotic metaphases of C. capillaris showing elongation of bivalents and fragmentation. TABLE 3 First Metaphask Counts from Single Buds op Plants of C. capillaris Showing the Amount of Non-Conjunction Observed Plant 3* 2*+2' r+4' [27H.21-4 63 9 Y . • 28H.A-7 X-strain < 129.169-3 84 20 62 78 4 [29. 169-3 45 89 1 27.12-23 153 1 28.142-11 66 4 64 University of California Publications in Agricultural Sciences [Vol. 6 29.169-3 of the X-strain and 28.142-11 of another strain were flower- ing in winter when capillaris does not thrive. The former showed a great number of irregularities while the latter was nearly normal in spite of the fact that the material was secured when the plant had almost completed its life and was beginning to die — circumstances which have been found to be associated with meiotic irregularities (Hakannson, 1926). This indicates that high percentage of irregu- larities cannot be ascribed to unsuitable environment and this is further upheld by the fact that the other plants of the X-strain flowered in the summer under essentially similar conditions and showed equalty wide variation in amount of irregularity. Root tips of 29.169-3, which was the most irregular of any plant studied at all extensively, showed a normal somatic chromosome complex. Tetrad studies showed different amounts of irregularities in the shape of micronuclei, microcytes, supernumerary cells, and diads in different plants of the X-strain, but no irregularities were observed in two plants of another strain (27.12-23, which showed almost regular I-M, table 3, and a sister plant not examined at I— M). Pollen studies gave results to be expected from the meiotic phenomena. While tectorum plants gave almost always 95 per cent or more good pollen, the X-strain of capillaris gave generally higher percentages of bad pollen than plants of tectorum and other strains of the same species, the percentage varying greatly from plant to plant. Table 4 gives various pollen counts from single heads of 12 plants of the X-strain and 3 plants of other strains. The second to the eleventh plants are selfed progeny of the first plant, the twelfth is a third- generation plant from the same source, while the other three represent other strains. All counts were made from healthy plants grown in the summer except 29.169-3 and 28.142-11, which flowered in the winter. With the exception of the third and fifth plants, counts made from different heads of the same plant on the same or different days showed little difference in percentage of good pollen, and even with them the difference was not very great. The various sister plants of the X-strain showed wide differences in the amount of good pollen, which in several cases was known to have been associated with corre- sponding differences in the amount of meiotic irregularities. Thus 27.12-23 (no. 14 in table 4) was almost completely regular at I-M (table 3) and had scarcely any bad pollen, plant 27H.21-4 (no. 6) had a few more I-M irregularities and had a higher percentage of 1930] Hotting she ad: Hybrids of Crepis capillaris and C. tectorum 65 TABLE 4 Pollen Counts from Various Flowers of Fifteen Plants of C. capillaris, Where Bracketed Together the Counts Were Made on the Same Day Plant Number of grains Bad Good Bad per cent 1 26.X-2 596 36 5.7 2 27H.8-2 [418 11 2.5 \365 17 4 4 3 27H.8-4 [359 47 11.6 \366 47 11.4 [403 110 21.4 \ 566 51 8.3 [420 39 8.5 (397 38 9.6 4 27H.8-12 ( 6 237 97.5 1 17 234 93.2 / 23 209 90.1 125 226 90.0 5 27H.8-13 [ 59 309 74.3 1 49 410 89.3 6 27H.21-4 672 114 14.5 7 27X-1 [434 63 12.7 1606 62 9.3 8 27X-3 [234 375 61.6 \ 94 170 64.4 9 27X-12 376 137 26.7 10 27X-13 (31 211 87.2 1 34 704 95.4 11 27X-14 567 144 20.2 12 29.169-3 197 446 69.4 13 27. 1817-5 almost all scarcely any 1- 14 27. 12-23 almost all scarcely any 1- 15 28. 142-10 488 18 3 5 bad pollen, and 29.169-3 (no. 12) was highly irregular at I— M and had a very high percentage of bad pollen. Plant 27H.8-12 (no. 4), which had the highest percentage of bad pollen, had the most irregu- lar tetrads of any plant examined ; an examination of a single slide showed many irregular I-M plates and only a few with the normal three bivalent chromosomes. Plants which were very irregular were completely, or almost completely, sterile. This probably indicates that gametes with abnormal chromosome complexes resulting from irregular meiosis usually fail to develop and that generally only normal gametes function. During the in- vestigation more than 2500 interspecific hybrids involving various 66 University of California Publications in Agricultural Sciences [Vol.6 plants of the X-strain of capillaris were obtained and, with the fol- lowing exceptions, capillaris plants appeared to have given normal hybrids with tectornm. The exceptions were: (1) a number of hybrids contained a diploid set of capillaris and a haploid set of tectornm, and (2) one hybrid examined contained the haploid complexes of the parental species and a single extra chromosome of capillaris. The first would indicate that there is a considerable number of female gametes with the somatic chromosome complex formed in capillaris and that they function, as had already been supposed from the occurrence of polyploids and triploid hybrids (Nawaschin, 19256, 1927). It was noted above that diads were seen occasionally and large pollen grains were observed rather often in some slides, indi- cating perhaps that somatic male gametes were also formed, although non-reduction was not actually observed. The second exception shows that female gametes with an extra chromosome resulting from meiotic irregularity may function occasionally, which has already been sup- posed from the occurrence of trisomies in this species (Nawaschin, 1926). MEIOSIS IN CAPILLARIS-TECTORUM F, HYBRIDS The outstanding characteristics of the meiosis of these F 1 hybrids are a variable number of bivalents and a variable behavior of uni- valents. Two groups of hybrids involving two different strains of tectornm were studied in this connection and I— M counts of the number of bivalents and univalents were made on seven plants of the first group and three plants of the second. The results are given in table 5 which shows the frequencies of all the possible combinations of bivalents and univalents observed in the various plants. Each line represents the results obtained from a single slide made from a single bud and includes all the cells in which the bivalents and univalents could be counted and distinguished from each other. The two middle columns contain the combinations hardest to distinguish and the actual proportion of these cells may have been somewhat higher than the counts represent. A striking difference in distribution between the first and second groups was observed as soon as plants of the second class were exam- ined. Unfortunately material had been preserved from only three plants of the second group since no such difference had been antici- pated. Each group included hybrids grown in two different seasons. 1930] H oiling she ad : Hybrids of Crepis capillaris and C. tectorum 67 TABLE 5 The Frequencies of the Various Combinations of Bivalents and Univalents in capillaris-tectorum F t Hybrids Involving Two Different tectorum Strains Parental tectorum strain Hybrid 3-+1' 2 '+3' r+5- 7' 27 H 17-1 37 37 7 27 H 17-2 34 49 15 3 27 H 17-4 19 15 6 1498 27 H 20-1 12 7 1 27 H 15-6 31 14 2 27 H 15-2 12 3 3 28 H 17-1 f 47 16 i Total 52 17 3 244 158 37 3 ( 27 H 19-5 32 44 44 45 1648 27 H 12-1 12 21 16 20 . 1 28 H 128-4/ 16 39 31 38 1 Total 45 66 53 54 105 170 144 157 Fig. I. C. capillaris-tectorum F„ late diaphase, showing respectively (1) two, (2) one, and (3) no bivalents. 68 University of California Publications in Agricultural Sciences [Vol. 6 The plants of the group involving the 1498 strain usually showed three bivalents and one univalent, fewer cases of two bivalents and three univalents, and occasionally one bivalent and five univalents, and rarely or never seven univalents. Some variation in the propor- tion of these classes occurred from plant to plant but the type was similar in all. The three hybrids involving the 1648 strain showed a very different distribution. In these plants each possible combina- tion was represented in approximately equal proportions, the com- bination of three bivalents and one univalent, which was most Fig. J. C. capillaris-tectorum F,, first meiotic metaphase, (1) three bivalents, (2) two bivalents, (1) one bivalent, (4—6) no bivalents. frequent in the first group, being least frequent here. There could be no doubt that for the plants examined a very different type of distribution of possible combinations characterized the two groups. Three PMC's at late diaphase representing the second, third, and fourth combinations are shown in figure I. Size differences can be seen but it is impossible to distinguish the various chromosomes. Cells at I-M showing all possible combinations of bivalents and univalents are shown in figure J. The bivalents resemble those in the parental species and are not constant in shape. The univalents usually lie away from the metaphase plate but one or more may lie beside the bivalents (fig. J, 2, 3). The cells containing seven unpaired chromo- 1930] Hollingshead: Hybrids of Crepis capillaris and C. tectorum 69 somes are especially striking. The univalents appear to be scattered throughout the cytoplasm and do not form a typical metaphase plate. Their arrangements frequently simulate anaphases (fig. J, 5, 6), but that these are not anaphases is shown by their association with cells at metaphase and by the fact that none yet show the split which is characteristic of anaphase chromosomes. Slightly later such figures Fig. K. C. capillaris-tectorum F„ first meiotic anaphases showing both segregation and division of univalents. as those shown in figure J, 5 and 6 will pass over into anaphase with- out any major change in the relative positions of the chromosomes. Occasionally a very unequal distribution of chromosomes, such as six at one pole and one at the other, was seen, but no certain case of seven univalents at one pole was observed. Figure J, 4 shows seven in a group nearer one pole than the other but the ones near the middle may move toward the other pole in the following anaphase. The kind of anaphase is apparently determined by the number of univalents and the positions they assumed at metaphase. Various 70 University of California Publications in Agricultural Sciences [Vol. 6 I-A arrangements are shown in figure K. If the univalents have lain on or near the plate at I-M they apparently divide, the halves going to either pole (fig. K, 2-5). If they were not on the plate they move toward the nearest pole with the dissociated bivalent partners (fig. K, 1, 4). Figure K, 1, shows a cell in which no univalents have divided and there are three split chromosomes at one end and four at the other. Figure K-2 shows one univalent dividing on the plate with three split chromosomes at each pole. The other anaphase figures Fig. L. C. capillaris-tcctorum F 1? first meiotic anaphases showing ' ' unclean ' ' division, fragmentation, and lagging chromosomes. Fig. M. C. capillaris-tectorum F„ (1) second meiotic prophase with very unequal nuclei, (2) micronucleus at interphase. doubtless represent cases in which less than three bivalents had been formed. Figure K, 5 shows a case of five splitting univalents on the plate, the highest number which was seen dividing at this stage. It may have arisen from a cell with one bivalent or from a cell in which all the chromosomes were unpaired and five of them lay on or near the plate. The former seems more likely. An "unclean" separation of partners of bivalents, which has been frequently described for other hybrids, and extreme attenuation of anaphase chromosomes sometimes resulting in fragmentation were seen occasionally in some slides and rather frequently in others. Figure L, 1 is a drawing of a I-T figure showing "unclean" separa- tion and fragmentation. In figure L, 2 a chromatin body which may 1930] Hollingshead: Hybrids of Crepis capillaris and C. tectorum 71 be either a chromosome or a fragment has been left in the cytoplasm and such a condition is not uncommon. Micronuclei are seen fre- quently in interphase (fig. M, 2). No case of a single nucleus at interphase was seen but instances of nuclei of very different size were occasionally observed. Figure M, 1 shows a case in which one nucleus in second prophase apparently consists of a single chromosome. ^ Fig. N. C. capillaris-tectorum F,, (1-5) second meiotic anaphases showing both division and segregation of univalents, (6) second telophase showing chromo- some elimination. Second divisions present a regular or irregular appearance de- pending on the nature of the preceding division. A first division in which the univalent chromosome or chromosomes did not divide but were included in the interphase nuclei apparently is followed by a regular second division in which all the chromosomes divide (fig. N, 1). A first division in which the univalent chromosome or chromo- 72 University of California Publications in Agricultural Sciences [Vol. 6 somes divided and the halves were incorporated in the interphase nuclei gives rise presumably to an anaphase with the univalent halves lagging on the spindle (fig. N, 2) and finally passing undivided to the poles. Figure N, 3 and 4, probably arose from interphases which con- tained micronuclei. In figure N, 3 at least one univalent half is passing undivided to a pole along with the halves of the bivalent part- ners. That the univalent chromosomes divide at either division is evident from the figures. However, a study of thirty clear second anaphases showed a total of fourteen chromosomes in every cell, indicating that the univalents divide only once during meiosis. An excessive drawing out of chromosomes at II-A similar to that de- scribed for I-A occurs occasionally, and fragmentation was evident in a few cases. In the cell shown in figure N, 4 there was only one II-M plate and it contained all but three chromosomes ; it presumably arose from an interphase with one large nucleus and one or more micronuclei. Figure N, 5 depicts a clear instance of a process which would give rise to somatic gametes. The seven divided chromosomes are of the second-division type and the cell was in a row of cells showing second- division figures. It could have arisen from a single interphase nucleus which resulted from a segregation of seven univalent chromosomes to one pole. Although such an occurrence was not seen, it has been pointed out above that very unequal segregations were observed and it is likely that all the chromosomes will occasionally pass to one pole. On the other hand, . it is possible that a restitution nucleus, such as that described by Rosenberg (1926), may have been formed following the first division, although no evidence of any such process was seen. It is evident from second-anaphase figures that nuclei with differ- ent numbers of chromosomes will sometimes occur and this is borne out by second-telophase figures where nuclei of various sizes may be seen. Figure N, 6 shows two pairs of nuclei of different sizes con- taining several chromosomes each and two chromosomes lying in the cytoplasm which would probably have given rise to micronuclei. The tetrads (fig. O), particularly from the hybrids involving the 1648 strain of tectorum, show frequent irregularities. Micronuclei and microcytes of various sizes are common and diads with and without micronuclei and microcytes are often found. It is easy to picture how some of the tetrads shown may have arisen from second anaphases, for instance figure 0, 1 from such a cell as shown in figure N, 1 ; figure O, 7, from figure N, 4, and figure O, 8, from figure N, 5. wao] H oiling she ad : Hybrids of Crepis capillaris and C. tectorum 73 Some variation occurred in the percentage of well stained pollen obtained from different plants, but it was low in all cases, usually less than 1 per cent. Two F x plants had a considerably higher per- centage (15 per cent and 21 per cent). One of these proved to be a triploid hybrid (below), the other was not examined cytologically. Considerable variation in pollen size occurred and large well stained grains, presumably arising through non-reduction, were found quite frequently. 9 W Fig. O. Tetrads of C. capillar -is-tectorum F^ The meiotic behavior of this hybrid differs from that described by Collins and Mann (1923) for Crepis setosa-C. capillaris and from that described by Nawaschin (1927) for C. capillaris-C . aspera F x hybrids in the fact that bivalents are formed. Each of these hybrids, like the capillaris-tectorum hybrids described above, is a hybrid of capillaris with a four-chromosome species. 74 University of California Publications in Agricultural Sciences [Vol. 6 The capillaris-setosa hybrid (Collins and Mann, 1923) was among the first examined in the genus and has been widely quoted in the literature as an example of complete lack of chromosome pairing. In later (unpublished) investigations, however, Mann Lesley found a variable number of bivalents in this hybrid. The later material involved different strains of both species. It was shown above that capillaris-tectorum hybrids involving different strains of tectorum exhibited a marked difference in the frequency of cells with the various possible combinations of univalents and bivalents. In one group of hybrids the cells with all the chromosomes unpaired were very frequent and especially striking. In the other group they were extremely rare and the occurrence of bivalents was the rule. It is probable that such a phenomenon may explain the difference between Mann Lesley's earlier and later observations. Possibly the earlier material had a predominance of cells with all chromosomes unpaired and those with bivalents were overlooked, while in the later material more cells had bivalents. In the capillaris-aspera hybrid described by Nawaschin (1927) no bivalents occurred and none, some, or all of the univalents divided at the first division, the division having been initiated at diaphase. If all the chromosomes divided at first division a diad was formed, other- wise the second division appeared to proceed normally. It is very possible that some of the diads found in the capillaris-tectorum hybrids of this paper were formed in this way, but as was pointed out above a different mode of diad formation was observed. The initiation of univalent division at diaphase was not observed in capillaris-tectorum hybrids. The meiotic behavior of the capillaris-tectorum hybrids is essen- tially similar to that described by Babcock and Clausen (1929) for three other interspecific hybrids in this genus. These hybrids involved only species with four chromosomes and each showed a variable num- ber of bivalents with a maximum of four, the highest possible number. 1930] H oiling she ad : Hybrids of Crepis capillaris and C. tectorum 75 TRIPLOID HYBRIDS (2 n capillaris, n tectorum) Attention was drawn to the first of these triploid hybrids, which occurred among a number of normal Fj hybrids, by its higher per- centage of good pollen and by its fertility, a considerable number of achenes per head having been formed while normal F 1 hybrids had none or rarely one or two achenes per head. The plant was then caged and a number of selfed seed obtained from which progeny to be described later were derived. When the next season's hybrids were grown some larger rosettes were set aside as being possible triploid hybrids and four of them proved to be so. Another plant which was selected at maturity, by its fertility proved on later examination to have been of the same chromosome constitution. Of the four of this group under observa- tion from the rosette stage, only one produced any seed and from it a selfed progeny was obtained which will be discussed later. The triploid hybrids resembled normal F x (Hollingshead, MS) but showed the effect of the extra haploid capillaris chromosome com- plex in a slightly stronger resemblance to that species. The somatic cells showed clearly two haploid sets of capillaris and one of tectorum (fig. P, 1). As was shown by Nawaschin (1927) for similar hybrids the D-chromosome of tectorum lacks the satellite here as it does in normal F x . The first meiotic division showed almost always three bivalents and four univalents at diaphase (fig. P, 2) and metaphase (fig. P, 3). Presumably the three bivalents are formed from the diploid capillaris complex and the four univalents are the tectorum chromosomes. It is worth noting that whereas capillaris and tectorum chromosomes pair in the F 1 hybrid the tectorum chromosomes here usually do not unite with the capillaris bivalents to form trivalents. Only rarely does one see six units at metaphase, one of which is probably a tri- valent (fig. P, 4). Rarely, too, only two bivalents were formed and six univalents were to be seen. This lack of pairing within a diploid capillaris complex was noted above in the pure species. At I-A the univalents segregate undivided or divide, the halves passing to either pole as shown in figure P, 5 and 6, their behavior having been determined apparently by their metaphase position, as 76 University of California Publications in Agricultural Sciences [Vol. 6 in normal F x hybrids. Lagging chromosomes at II-A were common and tetrads with microcytes and micronnclei frequent. Though no extensive examination of second-anaphase figures was made, several cells showed a total of twenty chromosomes at this stage and it was T r At Fig. P. Triploid C. capillaris-tcctorum hybrid, (1) somatic metaphase, (2) diaphase, (3) first meiotic metaphase showing three bivalents and four univalents, (4) first meiotic metaphase showing two bivalents, a trivalent, and three unival- ents, (5, 6) first meiotic anaphases showing both segregation and division of univalents. definitely established that certain chromosomes segregated wit bout division at second metaphase. In view of these observations and the behavior of ¥ x univalents it seems likely that univalent chromosomes 1930] Eollingshead: Hybrids of Crepis capillaris and C. tectorum 77 do not often, if ever, divide twice in this triploid hybrid. In spite of the fact that irregularities were more obvious in triploid hybrids than in most F x hybrids examined, the percentage of good pollen was much higher, averaging from 17.2 to 28.8 per cent, and, as has been noted, three of them were somewhat fertile. The meiotic behavior of the triploid hybrids, 2 n capUlaris-n aspera, described by Nawaschin (1927), showed three bivalents and four univalents but, unlike the capillaris-tectorum triploid hybrids, all the univalents divided at the first division, second divisions were regular, and almost always regular tetrads occurred. This would mean that the univalents divided at both divisions and that every gamete would have the somatic chromosome set. On this basis one would expect a high percentage of morphologically good pollen and practically com- plete fertility. Nevertheless very little good pollen was produced. No selfed progeny were secured by Nawaschin from his triploid hybrids. PROGENY OR TRIPLOID HYBRIDS The meiosis of the triploid hybrids would lead one to expect that each male gamete (and presumably female) would contain a haploid set of capillaris chromosomes and one to four tectorum chromosomes. If the segregation of the tectorum univalents were a random one, fifteen-sixteenths of the gametes would possess at least one tectorum chromosome in addition to a haploid capillaris set. If all gametes were equally viable the zygotes derived from them would possess a diploid set of capillaris and none to eight tectorum chromosomes, i.e., a range of chromosomes number from six to fourteen. Table 6 gives the actual distribution of chromosome numbers in plants from the two TABLE 6 The Actual Distribution of the Chromosomes Numbers of the Progenies of Two Triploid C. capillaris-tectorum Hybrids Compared with the Theoretical One Chromosome number 6 7 8 9 10 n 12 13 14 Progeny 1 Progeny 2 10 49* 5 7 2 2 1 3 1 11 1 2 1 2 3 1 Total 59 12 4 4 12 3 3 3 1 Theoretical 1 8 28 56 70 56 28 8 1 * Only nine of these plants were actually counted. The constitution of the others was inferred from the fact that they were morphologically typical capillaris plants. 78 University of California Publications in Agricultural Sciences [Vol. 6 selfed progenies compared with the theoretical one. The most out- standing feature is the difference between the actual and theoretical proportion of plants with six chromosomes only, i.e., with no tectornm chromosomes. They constitute more than half of one population by actual chromosome determination, and the same is doubtless true of the second population in which most plants which were morpholog- ically like pure capillaris were not examined cytologically, whereas the theoretical expectation is only 1 in 256. The distribution of the chromosome numbers of the plants which contained some tectornm chromosomes differs from the theoretical, the seven-chromosome class being too high in both populations. Were numbers alone taken into consideration one might postulate that gametes with tectornm chromosomes suffered in competition with those without, and those with one suffered least, thus giving rise to many zygotes with capillaris chromosomes only and a larger propor- tion with one tectornm chromosome than was expected. When the complexes of the plants were analyzed to determine exactly which chromosomes were present, however, and this was done for most plants, this explanation in its simplest form proved untenable. As expected, each plant did contain a diploid set of capillaris. Accord- ing to chance combinations of the gametes derived from random segregation of univalents, many plants should have contained at least one pair of tectornm chromosomes. Of the ten-chromosome plants, for example, 97 per cent would have at least one tectornm chromosome represented twice and only 3 per cent would contain the four different tectornm chromosomes. Of the twelve plants with ten chromosomes all had the four different tectornm chromosomes. Indeed no tectornm chromosome was found twice in a plant unless all four were present once. The results are explicable on the assumption that egg cells with most, or all, of the possible combinations of chromosomes may func- tion but that only those pollen grains function which have either a haploid capillaris set or a haploid set of both species, each of which would occur once in sixteen times as a result of random segregation of univalents. The high percentage of poor pollen accords well with the hypothesis that considerable pollen is non-functional, although the proportion of obviously bad pollen is not high enough to account for the postulated elimination. Probably all well stained grains cannot function. On this assumption, the constitutions of the female gametes which gave rise to particular plants may be determined. If the plant had 1930] H oiling shead : Hybrids of Crepis capillaris and C. tectorum 79 less than ten chromosomes, the gametic constitution of the egg from which it arose can be obtained by subtracting a haploid capillaris set from the whole chromosome complex. If the plant had ten chromo- somes, the female gamete contained either a haploid capillaris set or haploid sets of both species. If it had more than ten, the gametic constitution of the egg from which it arose can be determined by subtracting the haploid sets of the two species from the whole com- plex. On this basis the distribution of functioning female gametes with one to three tectorum chromosomes in addition to the haploid capillaris set proves to be : Number of tectorum chromosomes Frequency 15 It is impossible to determine the frequency of functioning female gametes with four tectorum chromosomes since pollen grains of the same constitution presumably function also. Since the two triploid hybrid progenies were apparently funda- mentally similar in the distribution of chromosome numbers from which these calculations were made, they were grouped together for this purpose. In one detail, however, they were quite different, the second having a much higher proportion of ten-chromosome plants; and the difference seems to be too high to be ascribed to the small number of plants in each population. According to the hypothesis outlined above this might mean that one parent had produced a higher proportion of gametes with four tectorum chromosomes but there is no further evidence for such a deduction. From those plants where it was possible to determine exactly which chromosomes were present, it was calculated that the distri- bution of various female gametic chromosome combinations had been as follows (using the chromosome designations as in fig. B) : Gametic constitution n capillar is +T A n capillaris-{-T B n capillaris +T D n capillar is +T A , T B n capillaris +T B , T D n capillaris+T B , T c n capillaris+T A , T B , T c n capillaris +T A , T B , T c , T D Frequency 6 3 1 2 1 2 1 At least 1 80 University of California Publications in Agricultural Sciences [Vol. 6 Besides the plants from which this distribution was calculated there occurred seven others each of which contained a chromosome which looked like T A with a part of one arm missing (fig. Q, 1). These plants each had a diploid set of capillaris, a haploid set of tectorum, and this peculiar chromosome, with or without part of another haploid set of tectorum which never included T A . The repeated occurrence of this chromosome, which may be a T A from which a part has been lost by fragmentation and elimination at meiosis, might indicate that the A-chromosome of tectorum has a tendency to fragment at a certain point. The situation recalls that found by Nawaschin (1927) in the plants of the progeny of a triploid Fig. Q. Somatic metaphases of two derivatives of a triploid hybrid, (1) a plant with eleven chromosomes, one of which (x) is a type not found in the parental species (see text), (2) the amphidiploid derivative. capillaris-aspera hybrid. Each had a typical A-chromosome of capillaris and one with a shorter arm than usual, which he inter- preted as showing that hybridization had affected the shape of the chromosome. In later progenies of the same type, however, he was unable to find these modified chromosomes (unpublished data) and now inclines to the fragmentation hypothesis. In two plants which had two D-chromosomes of tectorum one of these was satellited, the other was not. In all other cases each T D - chromosome lacked the satellite, as in the F t and the triploid hybrids. No explanation for the reappearance of the satellite was found. A number of possible combinations of tectorum chromosomes are missing. The number of plants examined is too small to draw any conclusion from this fact but differential viability among the plants which did have different combinations favors the possibility that some chromosome combinations are quite inviable. Each particular combination of tectorum chromosomes added to the capillaris complex had its peculiar effect on the morphology of 1930] Eollingshead : Hybrids of Crepis capillaris and C. tectorum 81 the plant. All plants with a diploid capillaris complex and part of a haploid tectorum set were less viable than plants with only capillaris chromosomes, and the plants with two tectorum chromosomes were usually less viable than those with one or three. Plate 2, a, shows a typical capillaris plant at the rosette stage and three others, each of which had a different tectorum chromosome added to the capillaris complex. T B was most injurious to viability. Plate 2, b, shows four plants with more than one tectorum chromosome added to the capil- laris complex. The combination of T A T B with capillaris was the least viable one obtained. Those which flowered under the writer's observation were sterile. Plants having ten chromosomes, with one exception, resembled the triploid hybrid from which they arose. The one exception was the single plant in the first population which had ten chromosomes. It was rather weak and sterile, and differed in habit and leaf shape from its parent. The chromosome complex showed a diploid set of capillaris, A-, B-, and C-chromosomes of tectorum and a satellited chromosome which could not be distinguished from the capillaris satellited chromosomes. Probably non-disjunction or non-conjunction in the capillaris set had given rise to an egg with two capillaris satellited chromosomes instead of the usual one. Plants with more than ten chromosomes formed vigorous rosettes. Only the first population was under the writer's observation during the whole life of the plants and the one plant with twelve chromo- somes never flowered. The other with eleven was sterile. Some of those in the second population were in flower when the writer had to discontinue the investigation, others were still at the rosette stage. They showed morphological differences but were more uniformly viable than those with fewer than ten chromosomes. The possibility of hybridization having played a part in the evolution of chromosome numbers within this genus has been sug- gested by Nawaschin (1925&) and Hollingshead and Babcock (1930). The plants just described, however, constitute evidence against the hypothesis which would explain the origin of a species with a higher chromosome number by the addition of a pair of chromosomes, from another species to the complex of an existing one following hybridiza- tion. As has been pointed out, the addition of a single chromosome of tectorum to the capillaris complex lessened viability and caused sterility. It does not necessarily follow, of course, that the same would hold good for derivatives of other interspecific hybrids within the genus. 82 University of California Publications in Agricultural Sciences [Vol. 6 THE AMPHIDIPLOID DERIVATIVE The nature of the progeny of the first triploid hybrid favored the possibility of obtaining a plant with a diploid chromosome complex of each species, and it was largely in the hope of obtaining such a plant that the second population was grown and examined. When such a fourteen-chromosome plant was obtained, disappointingly enough, although containing the expected chromosome constitution, it was somewhat spindly and weak compared with the stouter, though later, capiUaris and triploid hybrid plants of the same population (pi. 3). It produced no typical rosette but sent up a weak flower stalk very early. Many heads w r ere abnormal with greenish yellow split ligulae and no pollen, a phenomenon occasionally met with in F x hybrids. Some heads however, were normal in appearance and in other characters it resembled a normal capiUaris- tectorum ¥ 1 hybrid. Part of the weakness was doubtless due to its being grown in the winter, but this characteristic was too marked to be attributed to environment alone. TABLE 7 First Metaphase Counts from Three; Buds of the Amphidiploid C. capillaris-tectorum Hybrid 7' 6'+2' 5'+4' 4"+6' 6 6 10 2 7 9 2 2 4 1 Total 22 18 8 1 The somatic chromosome complex is shown in figure Q, 2. Each tectorum D-chromosome lacks its satellite. Meiotic divisions showed the expected seven pairs in less than half of the cells. Table 7 shows the frequency of the combinations of bivalents and univalents observed in three slides. Since the higher the number of units the more 'difficult it is to distinguish the exact number of bivalents and univalents, it is probable that the irregularities were more common than the table indicates. Figures R and S show first diaphase and metaphase stages illustrating the various combinations observed and one anaphase showing a regular distribution of bivalent partners. Tetrads showed occasional microcytes and micronuclei. One normal 1930] Hollingshead : Hybrids of Crepis capillaris and C. tectorum 83 head gave 32.4 per cent apparently good pollen. The high percentage of irregularity and the sterility is disappointing, for hopes had been entertained that this would constitute an experimental verification of an hypothesis which would account for some of the various chromo- some numbers of the species within the genus by hybridization fol- Fig. E. First meiotic divisions of the amphidiploid derivative, (1) diaphase with six bivalents and two univalents, (2) with seven bivalents, (3) metaphase with seven bivalents, (4) with six bivalents and two univalents. lowed by chromosome doubling (Hollingshead and Babcock, 1930). That the irregularities can be attributed wholly, or even largely, to environmental conditions seems unlikely since some capillaris plants growing under similar conditions had regular meiotic divisions. It seems very probable, however, that the meiotic irregularities are 84 University of California Publications in Agricultural Sciences [Vol. 6 expressions of the same phenomenon as that met with in the X-strain of capillaris which is the strain incorporated in this amphidiploid hybrid. Under the circumstances it would be premature to conclude that similar hybrids would always be weak, meiotically irregular, and sterile, or to abandon the hypothesis that such hybrids may have figured in the evolution of the chromosome numbers in the genus. Fig. S. (1, 2) First meiotic metaphases with five and four bivalents respec- tively, (3) regular first anaphase. Nevertheless, in this connection it is worth noting that an amphidiploid C. capillaris-C. dioscoridis hybrid obtained by M. Nawaschin (un- published data) was smaller than F x hybrids of these two species and was sterile. Amphidiploids (also called didiploids, allopolyploids, and tetra- ploid hybrids) which involve two species of the same genus have been described in Nicotiana (Clausen and Goodspeed, 1925, Eghis, 1927, and Rhybin, 1927), Rosa (Blackburn and Harrison, 1924), 1930] Soiling shead: Hybrids of Crepis capillaris and C. tectorum 85 Prunus (Crane and Darlington, 1927 ), 1 Saxifraga (Marsden- Jones and Turrill, 1928), Solatium (Jorgenson, 1928), Digitalis (Buxton and Newton, 1928), and Primula (Newton and Pellew, 1929). Simi- lar combinations which involve the chromosomes of species of different genera have been described in Aegilops-Triticum hybrids by Tscher- mak and Bleier (1926). and in Raphanus-Brassica hybrids by Kar- pechenko (1927). As pointed out by Newton and Pellew, there are two known methods by which these have arisen from normal F 1 hybrids — the doubling of chromosomes in the soma of the F x hybrid and the production of unreduced gametes which give rise to amphidi- ploids directly or to triploid hybrids which subsequently give rise to amphidiploids. The derivation of the Crepis amphidiploid is of interest in that it, like the Nicotiana of Eghis and Rhybin, arose from a triploid hybrid which was the result of a union of an unreduced gamete from a pure species with a normal gamete from another. Thus the formation of the amphidiploid can be traced to the occur- rence of non-reduction in Crepis capillaris. Amphidiploids generally have been at least as vigorous as the F t hybrids from which they arose. The spindly nature of the capillaris- tectorum amphidiploid is particularly difficult to understand in view of the fact that F t hybrids and triploid hybrids involving these species are vigorous and healthy, except when an F x hybrid receives a lethal factor from its tectorum parent (Hollingshead, MS). While the amphidiploids which have been examined meiotically have shown small percentages of irregularities, in no other case has as high a percentage as that which characterized the Crepis amphidiploid been reported. One is the more inclined to attribute the high percentage of irregularity of this amphidiploid to the capillaris portion of the amphidiploid chromosome complex than to any effect of the com- bination of the two specific chromosome garnitures. 1 Actually a hybrid between tetraploid and diploid species, which arose from an unreduced gamete of the latter, giving rise to a plant with four haploid com- plexes. 86 University of California Publications in Agricultural Sciences [Vol. 6 SUMMARY 1. Examination of somatic chromosomes of the F x hybrid of C. capillaris (w = 3) and C. tectorum (to = 4) corroborated Nawa- schin's observation that the satellite on one chromosome of tectorum was not to be found in the hybrid. Measurements of the various chromosomes indicated that in the reciprocal hybrids examined another tectorum chromosome was shorter than in the parental species. 2. Meiotic divisions in tectorum proceeded normally but plants of the X-strain of capillaris frequently exhibited irregularities in the form of non-conjunction and irregular distribution of chromosomes. In extreme cases scarcely any good pollen was found. 3. Variation in number of bivalents at first meiotic metaphase characterized all F 1 hybrids. In hybrids involving one strain of tectorum three bivalents and one univalent was the most frequent combination and seven univalents were very rare. In those involving a second strain the frequencies of the various possible combinations of bivalents and univalents were approximately equal with three bivalents and one univalent the least frequent combination. The sug- gestion is offered that some such difference in hybrids involving different strains of a particular species may explain why Collins and Mann at first found no pairing in C. capillaris-C '. setosa F 1 hybrids, and later Mann Lesley found as many as three bivalents in similar hybrids involving different strains. 4. The univalents in the capillaris— tectorum hybrids divided either at the first or second division, but not at both. The hybrids were almost completely sterile. 5. Several triploid hybrids which contained a diploid set of cap- illaris and a haploid set of tectorum were found among ordinary P, hybrids and selfed progenies were obtained from two of them. The capillaris chromosomes united to form three bivalents at meiosis leaving the tectorum chromosomes unpaired. The chromosome con- stitutions of the progeny indicated that only male gametes which had none, or all, the tectorum chromosomes functioned but that most, or all, of the female gametes could function. Thus plants were secured with only capillaris chromosomes and with various combinations of tectorum chromosomes added to a capillaris complex. With the 1930] E oiling shead: Hybrids of Crepis capillaris and C. tectorum 87 exception of derivatives of the parental constitution (triploid hybrids) those with tectorum chromosomes were less viable than those without. This constitutes evidence against a theory which would account for an increase in chromosome number during the evolution of the genus by the addition of a pair of chromosomes from another species follow- ing hybridization. 6. One amphidiploid derivative with diploid chromosome com- plexes of each species was secured. It was less vigorous than triploid hybrids of the same population but matured earlier. Meiotic irregu- larities were very frequent and a large proportion of the pollen was bad, and it proved to be completely sterile. No explanation for its lack of vigor was found, but it may be that the meiotic irregularities were introduced by capillaris which was of the frequently meiotically irregular X-strain. Under the circumstances it would be premature to infer that the lack of vigor and the meiotic irregularity of this plant was evidence against the hypothesis which would account for some of the chromosome numbers within the genus by the occurrence of amphidiploidy. > 88 University of California Publications in Agricultural Sciences [Vol. 6 LITERATURE CITED Babcock, E. B., and Clausen, J. 1929. Meiosis in two species and three hybrids of Crepis and its bearing on taxonomic relationship. Univ. Calif. Publ. Agr. Sei., 2:401-432. Belling, J. 1928. A working hypothesis for segmental interchange between homologous chromosomes in flowering plants. Univ. Calif. Publ. Bot., 14:283- 291. Blackburn, K. B., and Harrison, J. W. H. 1924. Genetic and cytological studies in hybrid roses. I. The origin of a fertile hexaploid form in the pimpinellifoliae-vellosae crosses. Brit. Jour. Exper. Biol., 1:557-570. Buxton, B. H., and Newton, W. C. F. 1928. Hybrids of Digitalis ambigua and Digitalis purpurea, their fertility and cytology. Jour. Gen., 19:269-279. Clausen, E. E., and Goodspeed, T. H. 1925. Interspecific hybridization in Nicotiana. II. A tetraploid glutinosa- tabacum hybrid, an experimental verification of Winge's hypothesis. Genetics, 10:278-284. Collins, J. L., and Mann, M. C. 1923. Interspecific hybrids in Crepis. II. A preliminary report on the results of hybridizing Crepis setosa Hall, with C. capillar is (L.) Wallr. and with C. biennis L. Genetics, 8:212-232. Crane, M. B., and Darlington, C. P. 1927. The origin of new forms in Eubus. I. Genetics, 8:241-278. Eghis, S. A. 1927. Experiments on interspecific hybridization in the genus Nicotiana. I. Hybridization between the species Nicotiana rustica L. and Nico- tiana tabacum L. Bui. Appl. Bot. (Leningrad), 17:153-189. (Eng- lish summary.) Hakannson, A. 1926. Zur Zytologie von Celsia und Verbascum. Lands. Univ. Arschrift, N.F., 21:1-47. HOLLINGSHEAD, L. A lethal factor in Crepis effective only in an interspecific hybrid. MS. Hollingshead, L., and Babcock, E. B. 1930. Chromosomes and phylogeny in Crepis. Univ. Calif. Publ. Agr. Sci., 6:1. JORGENSON, C. A. 1928. The experimental formation of heteroploid plants in the genus Solatium. Jour. Gen., 19:133-211. Karpechenko, G. P. 1927. The production of polyploid gametes in hybrids. Hereditas, 9:349-368. Kuhn, E. 1928. Zur Zytologie von Thalictrum. Jahrb. f. wissen. Bot., 68:382-430. 1930] Hotting she ad: Hybrids of Crepis capillaris and C. tectorum 89 Marsden-Jones, E. M., and Turrill, W. B. 1928. A tetraploid Saxifraga of known origin. Nature, 122:58. Nawaschin (Navashin) M. 1925a. Polyploid mutations in Crepis. Triploid and pentaploid mutants in Crepis capillaris. Genetics, 10:583-592. 1925&. Morphologische Kernstudien der Crepis Arten in Bezug auf die Art- bildung. Zeit. f. Zell. u. Mikr. Anat., 2:98-111. 1926. Variabilitat des Zellkerns bei Crepis Arten in Bezug auf die Artbildung. Zeit. f. Zell. u. Mikr. Anat., 4:171-215. 1927. tiber die Veranderung von Zahl und Form der Chromosomen infolge der Hybridisation. Zeit. f. Zell. u. Mikr. Anat., 6:195-233. Newton, W. C. F., and Pellew, C. 1929. Primula Tcewensis and its derivatives. Jour. Genetics, 20:405-467. Rosenberg, O. 1926-27. Die semiheterotypische Teilung und ihre Bedeutung fur die Ent- stehung verdoppelter Chromosomenzahlen. Hereditas, 8:305-338. Bhtbin, V. A. 1927. Polyploid hybrids of Nicotiana tabacum L. X Nicotiana rustica L. Bui. Appl. Bot. (Leningrad), 17:191-240. (English summary.) Taylor, W. R. 1925. Chromosome constrictions as distinguishing characteristics in plants. Am. Jour. Bot., 12:238-244. 1926. Chromosome morphology in Fritillaria, Alstroemeria, Silphiwm, and other genera. Am. Jour. Bot., 13:179-193. Tschermak, E., and Bleier, H. 1926. Tiber fruchtbare Aegilops — Weizenbastarde Ber. deut. Bot. Gesell., 44: 110-132. PLATE 1 Typical plants of C. teotorvm, F\ C. capillaris-tectorum, and C. capillaris at maturity. Loo] PLATE 2 Plants of the progeny of a triploid hybrid showing the effect of various chromosomes of C. tectorum added to the capillaris complex. [92] UNIV. CALIF. PUBL. AGR. SCI. VOL. 6 [HOLL1NGSHEAD] PLATE 2 PLATE 3 Triploid hybrid progeny. From left to right, C. capillaris, amphidiploid, and triploid hybrids. [94]