CYTOLOGICAL STUDIES OF TRIPLOID PROGENIES OF P O P U L U S T R E M U L A

BY H E L G E JOHNSSON INSTITUTE FOR BREEDING FOREST TREES, SVALOF, SWEDEN

WO years ago the writer submitted some data concerning the T occurrence of triploid races of Populus tremula, crosses between diploid and triploid races being also dealt with (JOHNSSON, 1940). It was then shown that nine different clones of triploid aspen are known in Sweden. Both male and female clones occur. The chromosome number of P. tremula is x = 19. Meiosis in the triploids *shows at IM trivalents, bivalents and univalents and gives rise to gametes with chromosome numbers varying between 19 and 38. Further, unreduced gametes with n=57 are formed. Crosses between diploid and tri- ploid races are noteworthy in that individuals with aneuploid chro- mosome numbers are realized in high frequency, in which respect Populus tremula differs markedly from nearly all species in which 2x-3x crosses have been investigated. Crosses in both directions gave the same results, thus showing that the pollen grains are not more susceptible to numerically unbalanced chromosome numbers than the embryo-sacs. These studies have, however, been extended to embrace also crosses triploid X triploid, the results of which are submitted as a supplement to the earlier account of crosses diploid X triploid.

Diploid X triploid crosses. - As there is now available a far larger material of the combination diploid X triploid than that published in 1940, it may be justifiable to submit an analysis of the material as a whole, especially as the chromosome numbers have been determined for in all no fewer than 1,665 F , individuals. This analysis is given in the form of a diagram, Fig. 1, in which the broken line indicates the percentage distribution of F , individuals with different chromosome numbers from 47 different crosses diploid X triploid and reciprocally. This curve agrees almost entirely with that recorded in 1940 and based on a considerably smaller material. The only difference is that the curve has now received a more even course, a number of random irregularities having been eliminated. The mean of the chromosome numbers of 1,635 individuals (the tetraploid and approximately tetra- ploid have dot been included) is 46,7 as compared with 46,s for 617 individuals in 1940.

TRIPLOID PROGENIES OF POPULUS TREMULA 307

Triploid >( triploid crosses. - Four crosses between triploid and triploid aspen have been made. The fertility of the crosses is low both as compared with crosses between diploids and as compared with crosses in which only one of the parents is triploid. The fertility of the crosses and the germinability of the seed were as follows:

Number of crosses . . . . . . . . . . . . . . . . 18 5 4 Type of cross 2x >< 2x 3x >( 2x 3x x 3x

seeds per capsule . . . . . 9,g 433 2,4 ))

Gcrtninability of seed . . . . . . . . . . . . 94 % 45 9; 48 ”/. In this comparison the triploid parents in the 5 crosses 3x X 2x

are the same triploid 9-individuals as those employed in the 3x X 3x crosses. The low seed setting in the cross triploid X diploid as com- pared with diploid >(; diploid crosses must be due to a great many of the ovules of the diploid aborting, either as the result of the lethal constitution of the haploid (n-) egg-cell or on account OP diploid (2n-) lethal combinations arising at fertilization. The further reduction of the seed setting occurring in the cross triploid X triploid must be due to an increased frequency of lethal diploid (2n-) ‘combinations arising on fertilization.

Chromosome numbers o f the triploid offspring. - The chromo- some numbers have been determined for 308 individuals from the four triploid progenies, the following distributions being obtained :

2n = . . . . . . . . . . . . . . . 38 39 40 41 42 43 44 45 46 47 Number of individuals 17 3 2 4 5 6 1 5 10 11 2n = . . . . . . . . . . . . . . . 48 49 50 51 52 53 54 55 56 57 Number of individuals 11 8 11 4 11 13 12 23 21 56 2 n = . . . . . . . . . . . . . . . 58 59 60 61 62 63 64 65 66 67 Number of individuals 13 15 3 5 6 9 1 8 3 0 2n = . . . . . . . . . . . . . . . 68 69 70 7 1 72 73 74 75 76 Number of individuals 3 1 0 3 0 1 0 1 2 Total308

In the diagram, Fig. 1, a corresponding curve has been drawn (the unbroken line) after conversion to percentage of the total number of individuals. The most characteristic features of the curve are that practically all theoretically possible chromosome numbers are realized, and that there is a pronounced maximum for the triploid number, n=57. This maximum corresponds to the class which in respect of the chromosome numbers of the gametes (JOHNSSON, 1940) should be most numerously represented if no special selective mechanism is

I9 18. 17 16.

15,

I

-&-*/

--- 58 59 60 61

62 63 64 65 66 67 68 69 70 7l 72 73

74 75

- .

Fig. 1.

Diagram

showing the distribution of

chromosom

e numbers in the progeny after

the cross diploid X

triploid (- - - -)

and triploid x triploid Populus frem

ula ( ).

12

2

11 $

lo-\

2

-I -;

T-\

TRIPLOID PROGENIES OF POPULUS TREMULA 309

present. With regard to the theoretically probable distribution, niore- over, the diploid number, 2n = 38, is enormously over-represented, like the immediately following classes. Even the tetraploid number, n = 76, and the immediately lower classes must be considered to be over- represented. Some suggestion of a maximum is present also in the classes midway between the diploid and the triploid numbers for n = 45-50. There is an under-representation, however, within the nearest classes on either side of 2n=57. Further, the number of individuals between 2x and 3x is considerably greater (52,i 7; of the total number) than between 3x and 4x (23,3 % of the total number).

The chromosome number of the triploid progenies agrees entirely with that found in crossing triploid X diploid, the feature peculiar to the aspen that aneuploid individuals occur in a high frequency, owing to the fact that embryo-sacs and pollen-grains with aneuploid haploid numbers as well as embryos with aneuploid diploid numbers are viable.

The vitality of euploids and aneuploids. - The great majority of all individuals with aneuploid chromosome numbers possess a poor growth and have the appearance of standing on the border-line OF viability. Especially weak are such plants as possess more than 57 chro- mosomes. Still, one comes across an occasional individual with an- euploid chromosome number that shows good growth. For the pure euploid numbers an experiment has been arranged to throw light on their relative vitality. In this experiment a diploid, a triploid and a telraploid individual were propagated by root-cuttings. Ten 1-year-old plants of each clone were planted at random in a plot and were cut down to the surface of the ground on being planted. The diploid and triploid clones originate from wild stocks. There is therefore reason to assume that the triploid clone has arisen from a diploid via an unreduced gamete (cf. MUNTZING, 1936). The tetraploid clone originates from an individual that had arisen in a cross between the triploid of the experiment and a diploid aspen and must therefore be indebted to an unreduced gamete in the triploid for its origin. The development during the first summer is illustrated by the following mean measurements:

I.ength of stomata Plant height Diameter of stem rel. units cm. mm.

Diploid clone . . . . . . . . . . . . . 11,os 66,6 4,iio Triploid clone . . . . . . . . . . . . 13,iO 93,s 6,ti5

Tetraploid clone . . . . . . . . . . 14,411 82,2 7 , lO

31 0 HELGE JOHNSSON

The triploid and tetraploid clones have grown considerably faster than the diploid. However, it may be questioned whether the diploid contained in the experiment is a good representative of the average of the diploids. Probably many diploid clones occur with a better growth than the diploid of the experiment. The tetraploid clone, on the other hand, originates from the most vigorous individual of all the 34 tetra- ploid individuals. These 34 tetraploid individuals, 32 of which were obtained in crosses diploid X triploid, and 2 in crosses between triploid and triploid, possess a very variable vitality. The majority are very weak, and not less than 19 of the 34 died before 2 years of age. Only 3-4 give the impression of possessing full vitality. As triploids origin- nting from diploids by restitution nuclei and thus possessing three complete genomes are apparently superior to diploids in vegetative development, it seems as if triploidy is the optimal degree of polyploidy in P. fremula. Tetraploidy, on the other hand, seems as a rule to be associated with weak to languishing vegetative development. Only in exceptional cases do tetraploid plants occur with good vitality. The triploid plants of the cross-progenies are, in fact, almost without ex- ception more or less dwarfish and feeble in growth, in contrast to the triploids with gigas characters emanating from diploids by restitution nuclei. This is also readily explained by the fact that the triploids of the cross-progenies can extremely seldom possess three complete genomes, being in fact aneuploids despite the euploid chromosome number, the chromosome complement of which is characterized by sub- or supernumerariness for certain chromosome kinds, although the sum of the chromosomes is 57.

Discussion. - Of triploids in general it holds good that the chro- mosome numbers of their progeny do not agree with the chromosome numbers of their newly formed gametes, but, owing to a strong selection of gametes and embryos, individuals with aneuploid numbers are more or less completely absent. As a rule, the progenies both after self-fertilization and after crosses with diploids consist of diploids and individuals having one or two supernumerary chromosomes (e. g. Datura; BLAKESLEE and BELLING, 1934). In Allium schoenoprasum, LEVAN (1936) found that after self-fertilization the offspring of tri- ploids in the great majority of cases consist of tetraploids or individuals with one or two chromosomes less than the tetraploid number. In Populus frernula, on the other hand, the distribution of the chromosome numbers shows a close agreement with the probable distribution of gametes with different chromosome numbers. Also here, however,

TRIPLOID PROGENIES OF POPULUS TREMULA 311

there is a distinct over-representation of individuals with the diploid number in diploid X triploid crosses as well as in triploid X triploid. On the other hand, when diploid is crossed to triploid the theoretically

, do not coincide with most numerous chromosome numbers,

any euploid chromosome number. The maximum of the curve (Fig. l ) , which represents the distribution of chromosome numbers in the progeny, is also less pronounced. In a cross of triploid X triploid, however, the theoretically most numerous chromosome-number class,

38 + 57 2

57 + 57 , coincides with the euploid number 57, and the maximum of 2

the curve is also very marked. For the tetraploid number, too, there is a certain amount of over-representation. It is evident, therefore, that a positive selection value appears for whole multiples of the basic number 19.

Agreeing with P . tremula in regard to the viability of the aneuploids is Pyrus malus. Here, though, the chromosome numbers of the tri- ploid progenies seem to agree entirely with the probable random dis- tribution without any over-representation of even diploid individuals (WANSCHER, 1939). The explanation of the high frequency of an- euploids in P . tremula and Pyrus malus is probably the same, viz. that both Populus with n = 19 and Pyrus with n = 17 are secondary poly- ploids (JOHNSON, 1940), an explanation that has been accepted for Pyrus on other grounds by DARLINGTON and MOFFET (1930).

In the cross-progeny triploid X triploid P. tremula plants with chro- mosome numbers between 2n=38 and 57 are far more numerous than between 2n = 57 and 2n = 76 (see Fig. 1) . The cause of this can scarcely be an elimination of chromosomes at meiosis. In that case the maximum would have been displaced from 2n = 57 towards 2n = 38. Nor in all probability is the cause to be sought in an abortion

of gametes with a higher chromosome number than - for in a

37 diploid X triploid cross the interval between 2n = ~ -k 19 and 2n = 57 2 is well represented. The most probable explanation appears to be that embryos with 2n> 57 are aborted to a great extent. The lower vitality for aneuploids with 2n between 57 and 76 than for aneuploids between 38 and 57 is in good agreement with the fact that as a rule the tetra- ploid plants are not very vital. It is worthy of note that no individual has been encountered with 2n >76. In view of the relatively high

57 2 ’

312 HELGE JOHNSSON

frequency of unreduced gametes that is found among the triploids and that in crosses diploid X triploid results in 76-chromosome plants, plants with 2n>76 ought not ta be so very rare in the progeny after crossing triploid X triploid, provided they are viable.

It would therefore seem as if the optimal degree of polyploidy in Populus trernula is triploidy, and that tetraploidy constitutes the limit of vitality. In agreement with this is the fact that no tetraploid species or spontaneous race is known within the genus Populus.

LITERATURE CITED.

1. BLAKESLEE, A. F. and BELLING, J. 1934. Chromosomal mutations in the Jimson weed, Datum stramonium. - Journ. of Hered. 15.

2. DARLINGTON, C. D. and MOFFET, A. A. 1930. Primary and secondary chromo- some balance in Pyrus. - Journ. of Gen. Vol. 22.

3. JOHNSON, H. 1940. Cytological studies of diploid and triploid Populus tremula and of crosses between them. - Hereditas XXVI.

4. LEVAN, A. 1936. Zytologische Studien an Allium schoenoprasum. - Hereditas XXII.

5. MUNTZING, A. 1936. The chromosomes of a giant Populus tremula. - Hereditns XXI.

6. WANSCHER, J. H. 1939. Contribution to the cytology and life history of apple and pear. - Den kongl. Vet.- og Landbohejskoles Aarsskrift 1939. Kebeii- haw.