Jos UNIVERSITY OF CALIFORNIA PUBLICATIONS s ‘a ae BOTANY _ Vol. 5, No. 17, pp. 457-582, 2 figures in text, plates 55-85 April 14, 1922 eS A EAT TE INHERITANCE IN NICOTIANA TABACUM A REPORT ON THE RESULTS OF CROSSING ee CERTAIN VARIETIES ea WILLIAM ALBERT SETCHELL ; THOMAS HARPER GOODSPEED : ee

AND ROY ELWOOD CLAUSEN

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Studies in Nicotiana. “I, by William Albert Setchell. Pp. 1-86. Saeniihieg” BSS) Si iets ened i dane a ete a Ee We ERS Peal Me MeeR Ree CN aNee estan Hore dn Us 8 $1.25

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Quantitative Studies of Inheritance in Nicottana Hybrids. I, by Thomas Harper Goodspeed. Pp. 87-168, plates 1-28. December, 1912...

Quantitative Studies of Inheritance in Nicotiana ote aoe by Thomas . Harper Goodspeed. Pp. 169-188, plates 29-34. January, 1913 220... ,

. On the Partial Sterility of Nicotiana Hybrids made with : enue as Ss

Parent, by Thomas Harper Goodspeed. Pp, 189-198. “March, 1913~_

. Notes on the Germination of Tobacco Seed. I, “by Thomas Harper Good-

Speed.2 cP ps199-222- May, LORS ose Sa a eee, oath an esata wd Sea ese Ly

. Quantitative Studies of Inheritance in Nicotiana Hybrids. 1, by Thomas

Harper Goodspéed. ~ Pp; 223-231. “April,/1915 222

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Hybrids, by: 7. H. Goodspeed and J. N. Kendall. Pp. 293-299. November, ~

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- Phe Nature of the F, Species Hybrids between Nicotiana sylvestris and-

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Varieties of Nicotiana Tabacum, with Special Reference to the Conception -

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to Nicotiana, by John N. Kendall. Pp. 347-428, 10 text figures, plates 49- 535 <Mrarch, LOR8 ooo a he chee acorn ep tweepe nea pee Sree Controlled Pollination in Nicotiana, by Thomas Harper Goodspeed and = Davidson. Pp. 429-434. August, 1918 oes... ccs. cccek eee thee teen cnn lane ante rseee

, An Apparatus for Flower Measurement, by T. H. Goodspeed and BE. E. Clausen. Pp. 435-437, plate 54, 1 figure in text. September, 1918 maneae |

Note on the Effects of Hluminating Gas and Its Constituents in Causing.

Abscission of Flowers in Nicotiana and Citrus, by T. H. Goodspeed, J. M.-

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Inheritance in Nicotiana Tabacum. I,A Report on the Resuits of Crossing :

Certain Varieties, by William Albert Setchell, Thomas Harper Goodspeed,

and Roy Elwood Clausen: Pp. 457-582, 2. figures in text, plates 55-86. -

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Vol. 5, No. 17, pp. 457-582, 2 figures in text, plates 55-85

Ill.

IV.

{) A | : _)') UNIVERSITY OF CALIFORNIA PUBLICATIONS ; . BH IN Cb. BOTANY

INHERITANCE IN NICOTIANA TABACUM

i A REPORT ON THE RESULTS OF CROSSING CERTAIN VARIETIES > BY

WILLIAM ALBERT SHETCHELL, THOMAS HARPER. GOODSPEED, AND ROY ELWOOD CLAUSEN

April 14, 1922

¥

: CONTENTS PAGE

Introductonyy. wee... Mander: ee ah .. 458 Plan. of themwotkee:.......get.-.--. Peace ee .. 466 Angustifolia-macrophylla series............. .. . 462 1. Parents of the angustifolia-macrophylla series . 463 2. F, of the angustifolia-macrophylla series........ . 467 3. F. of the angustifolia-macrophylla series.. . 469 4. F; and subsequent generations of the Pe aiecmacropls ylla series...... 472 a. STENOPHYLLA derivatives.......... . 475

b. Latrroiia derivatives................ . 476

c. LANCEOLATA derivatives............ . 478

d. Loriro.ta derivatives . 478

e. AURICULATA derivatives........ Ps Tce , ine . 479

Ps. SESSHUUR OMAN G OM VATULV CSMAMMMN LS <x. .cc.d.0:/<uetee..:cesuseetdbes cy ssesde vader . 480

5. Summary of flower color observations in F, and paneeeent beneeations 482 6. Later sowings of F, and F; of the angustifolia-macrophylla series.. .. 483 7. Crosses of derivatives with the parents.......0..0.....:0ccccceeseeeeeee . 487 8. Discussion of results of the angustifolia-macrophylla series ................... _ 490 Calycina-wirginica SOMES; -.< eee cee ee oe ene eee, .. 494 1: Parents: of the-calycina-vimgunicaiseliesin seta. on nae neti . 494 2. F, of the calycina-virginica series.. . 496 3. F, of the calycina-virginica series................ Pa ae 497 4. F; and subsequent generations of the aplleatrey virginica series. . 499 5. Discussion of results of the a virginica series ............. 504 Alba-macrophylla series. BAAS . 504 1. Parents of the Gibeemaerophylan series... . 504 2. F, of the alba-macrophylla series................. .. 505 3. F, of the alba-macrophylla series................ ». 505 4. F; and subsequent generations of the alba- Piero phate series.. . O07 5. Discussion of results of the alba-macrophylla series ........ . 510

458 Umversity of California Publications in Botany [Vou. 5

PAGE

VI. Generalionclusionse 2) et o..: eee 510 1. Origin and interrelationships of varieties of Tabacwm.............00000000.-.---. 510

2. Methodology of Mendelian analysis in Tabacwme .........cccccccccecccecccecececececsen 513

3. Mendelianthereditivsimfabacwm.....cemees0. 2.) 516

VIL. Summarys siege. Beteec eee ete cacnes ss oot eS peak <1. 520 Iiterature cited!)....-...see eee Aca Oa SCC ERM sei ccensyte: ee 520 Explanation‘of plates: ea) ets... eese.....c. cee aera: ee 522

I. INTRODUCTORY

The inception of the work on the various species of Nicotiana grown and bred in the University of California Botanical Garden has already been sketched in a previous number of this series (cf. Setchell, 1912). As stated there, the original intention was to assemble a collection of tobacco plants simply as a portion of the outfit of the Botanical Garden for general instruction and display. So great was the variety and evident misapplication of the names under which the seeds were received, however, that it seemed advisable to attempt to determine, as definitely as possible, the status of each plant.

In this connection, the work of Comes, in particular, came under consideration and especially his views as to the origin and interrela- tionships of the various cultivated forms belonging to the T’abacum group. Comes (1899, p. 4 and elsewhere) regards the numerous eulti- vated forms of tobacco as having originated in various ways from certain fundamental varieties. He estimated that there are six of these fundamental varieties of Tabacum, and he supposed the large number of various and seemingly more or less intergrading forms to have arisen through the influence of the forces of acclimatization, adaptation, hybridization, and selection. Of these, undoubtedly, the greater variations have been produced and perpetuated, according to the ideas of Comes, through hybridization and selection. In his mono- graph (1899) and in his later more exhaustive treatise (1905), Comes has attempted to estimate just which of his six fundamental varieties of Tabacum have codperated in producing each one of the cultivated ‘‘races’’ so far as known to him.

The statements of Comes as regards the constitution of his various races seem to have been based on the results of morphological study rather than upon breeding analysis. The advisability occurred to the senior author of attempting to test Comes’ hypothesis by selecting varieties seemingly fundamental in type, and through hybridization

1922 | Setchell-Goodspeed-Clausen: Nicotiana Tabacum 459

and selection attempting to secure constant races exhibiting various recombinations of the parental characters. The work thus conceived has been carried out in detail in a certain few but seemingly charac- teristic cases. Several different crosses were made in 1909, the first filial generations were grown in 1910, and each year since that time has seen successive filial generations in the field.

Although the Nicotrana investigations were originally designed to attack experimentally a comparatively simple and definite problem, they have since been greatly amplified in scope. At the present time three rather distinct lines of investigation are actively in progress, viz.,

1. Mendelian inheritance in N. Tabacum.

2. Inheritance of quantitative characters.

3. Studies of interspecific hybrids. The recent appearance of bud variations in hybrid lines favorable for an analytie study of that phenomenon has resulted in the addition of another research project. Now it has been found that, although seemingly distinct, progress in these separate lines of investigation is more or less interdependent. In particular it has been found that certain of the phenomena exhibited in interspecific hybrid populations from crosses between NV. sylvestris and varieties of N. Tabacum require for satisfactory analysis and explanation an accurate and detailed knowledge of the Mendelian differences which exist among the par- ticular varieties of N. Tabacum that have been used in those investi- gations. Accordingly in later years these studies of hybrids between varieties of N. Tabacum, originally designed merely to test experi- mentally the interrelationships existing among such varieties, have been directed toward a specific Mendelian analysis of the germinal differences existing in a selected set of varieties. ;

With this change in emphasis has come a full appreciation of the difficulties of Mendelian studies in N. Tabacum. It has been very evident that, for the most part, the character differences among varie- ties of N. Tabacum do not rest upon a simple genetic basis; on the contrary, they often depend upon very complex and involved Men- delian differences; so that in segregating populations it is often im- possible to demonstrate the existence of definite, discontinuous char- acter classes. Not uncommonly the members of such populations may be arranged in series connecting by imperceptible differences the most extreme character expressions in the population.

But although complex intergrading segregation has often been observed in F’,, it has not been found that such complex segregation

460 University of California Publications in Botany [ Vou. 5

persists in subsequent generations in the hybrid lines. On the con- trary, it has been found, as will become evident in a study of the ex- perimental material communicated herewith, that a great simplification occurs in the segregation in F,, and subsequent generations, and that continuous segregation gives way to discontinuous just as might be expected from Mendelian theory. By observing the segregation in the consecutive generations of hybrid lines which have become homo- zygous in most of their loci through self-fertilization, it is possible to obtain some idea of the Mendelian factor pairs involved in the char- acter contrasts and of their relations to one another. It has also proved possible by a few years of self-fertilization to establish stable lines representing recombinations of parental characters. By investigating the interrelations among such stable derivative lines, which obviously should differ in fewer factors from one another and from the original parental varieties than the parental varieties differ from each other, it would seem possible to develop an indirect mode of attack by which the Mendelian analysis could be refined to any desired extent. The original plan of the investigation, therefore, having as its purpose a demonstration of the possibility of securing by hybridization stable derivative lines representing recombinatons of characters contained in the parents and comparable to the numerous existing varieties of NV. Tabacum, has been diverted into a detailed study of Mendelian differ- ences among a typical set of N. Tabacum varieties.

Il. PLAN OF THE WORK

In the introductory paper the senior author has diseussed the fun- damental types of N. Tabacum, and as indicated there, has expressed a preference for selecting some five fundamental varieties, or species, as representing the basal morphological elements found, or seemingly to be detected, in cultivated races of N. Tabacum. There is no neces- sity for discussing further, at present, the reasons for preferring the particular types selected by us as against those of either Comes or Anastasia (1906), since the fundamental conceptions agree sufficiently well and the important thing has been to make a beginning in experi- mentation by using varieties which present seemingly fundamentally different character complexes in most characteristic form in plants breeding true to type in the pure line. Certain reasons for selecting a particular type or types will be discussed in connection with the

1922 Setchell-Goodspeed-Clausen: Nicotiana Tabacum 461

consideration of the various crosses. Besides the ‘‘fundamental’’ types, there have been selected for crossing certain other types, possibly fundamental, or in some eases derivative, which have been employed for testing the inheritance of some particular character or group of characters. All of these have been described in the first paper of this volume.

The taxonomic problems in N. Tabacum do not appear to differ from those presented by many other species of cultivated plants. Barley, maize, oats, rice, wheat, among others, exhibit a similar diver- sity of forms with more or less obvious class ee tinctions: In these as in NV. T'abacum it appears to be an easy task to shuffle and recombine characters indefinitely. Clearly there can be no segregation of forms into distinct species on genetic grounds; the basis of speciation, if any, must depend either upon convenience merely or what amounts to practically the same thing, upon elevation of certain Mendelian char- acter contrasts to a higher rank in classification than others. Since

the taxonomic problem, therefore, is not strictly a genetic one, it seems best to follow general usage in this respect, eerring all the poly- morphie assemblage of forms to the one species NV. T'abacum, and re- garding the several races included thereunder as varieties of equal rank.

The varieties employed in this series of investigations are: WN. Tabacum var. alba, U. C. B. G. 30/06, previously described by Setchell as “‘White’’ Tobacco; NV. Tabacum var. angustifolia, U. C. B. G. 68/07, previously deseribed by Setchell as N. angustifolia; N. Tabacum var. calycina, U. C. B. G. 110/05; N. Tabacum var. macrophylla, U. C. B. G. 22/07; and N. Tabacum var. virginica, U. C. B. G. 78/05, previously described by Setchell as N. Tabacum ‘‘Maryland.’’ In each instance the University of California Botanical Garden (U. C. B. G.) number contains in the numerator the accession number of the year given in the denominator. The varieties have in the majority of cases been grown in pure lines from the date of their receipt. In order to avoid needlessly encumbering the text with scientific names, the varieties “mentioned above will be referred to by their varietal designations only, and when reference is made to the whole group the species name Tabacum will be used alone.

Three series of cultures are described in the present article: the angustifolia-macrophylla series, which has been derived from reciprocal crosses of angustifolia and macrophylla; the calycina-virginica series, derived in the same way from calycina and virginica; and the alba-

462 University of California Publications in Botany [ Vou. 5

macrophylla series, from alba and macrophylla. In the course of the investigations other crosses were made between different varieties of Tabacum and to a limited extent between other species of Nicotiana; but the principal attention has been paid to the three crosses noted above, and they and their progenies alone will be considered in the present paper. It may be said at this point that the different varieties of Tabacum cross readily with one another, giving an abundance of good viable seed. The hybrids are uniformly self-fertile.

The methods of hybridization used need not be considered here, because they have been deseribed in detail by Goodspeed (1912) else- where in this series. The particular refinements of technique which must be employed in sowing the seed, on account of its very small size, have also been there described. It might be well to state, however, that the most refined methods doubtless will not prevent the occasional appearance of a stray plant in the cultures. The danger of contami- nation arises not only during the sowing of the seed, but also when the bags are placed over the unopened buds. It is very easy to include a few stray seeds under the bag, for their small size makes it almost impossible to detect them in the coarse, sticky indumentum of the plant. In spite of these obvious difficulties, however, the number of plants that have certainly been strays has been very small. Their rare occurrence indicates clearly that the technique employed has been very successful.

III. ANGUSTIFOLIA-MACROPHYLLA SERIES

This series has received the most attention since the parents are so distinctly different, and the results have consequently been more complex than those which have followed the crossing of any other pair of Tabacum varieties. As will be demonstrated below, F, seemed at first hopeless in its variety of segregation. Later generations, however, exhibited so much less, or so little variety in their segregation products that it was easy to obtain new permanent combinations of characters or ‘‘fixations.’’ Certain of its segregants have been followed out to F,, and have also been crossed back on the parents which they most closely resembled.

Six successful crosses were made. Of these H, and H, had macro- phylla for the male and angustifolia for the female parent, while H,, H,, H,;, and H,, were reciprocal crosses. As a matter of convenience

1922] Setchell-Goodspeed-Clausen: Nicotiana Tabacum 463

the generations later than F, were grown only from H, and H,, the larger number from H,. The predominance of H, in the later families selected for the continuation of the work was not, however, due to any especially different behavior evidenced in that particular series.

1. PARENTS OF THE ANGUSTIFOLIA-MACROPHYLLA SERIES

By selecting angustifolia and macrophylla for crossing, two varie- ties were obtained which resemble each other in height and general habit, but which differ strikingly in leaf and flower characters. The differences are sufficiently great to lead one to regard them as belong- ing to different species; in fact, all five Tabacum varieties selected by us as possibly fundamental differ sufficiently among themselves to be regarded as species in the Tabacum section rather than as varieties. It is not our intention, however, to emphasize this point, since any discussion would of necessity lead to a general survey of all the known varieties and races at present included under Tabacum. If, however, these five, viz., angustifolia, macrophylla, ‘Cav ie,” Maryland, and ““Brazilian’’ (ef. Setehell, loc. cit.) could be considered by themselves as wild plants, it seems to us that any taxonomist of the present day would certainly award to each of them thejank of a separate species. These considerations should be borne in mind in estimating the signifi- eance of the results obtained through crossing.

Angustifolia, U. C. B. G? 68/07, is a variety which has long been known and which is répresented in our breeding experiments by a pure line very closely approximating the type. It has been figured and discussed by one of us (Setchell, loc. cit., p. 9, pl. 7). The photo- graph given there is of a young plant just coming into flower and consequently does not represent the habit of the plant in full blossom or in fruit, after the full number of laterals is developed. A plant in the height of its vigor is represented in plate 55, figure 1. In stature angustifolia belongs to the low corymbose group of Tabacum varieties, which also ineludes the forms bred in the University of Cali- fornia Botanical Garden under the names calycina and macrophylla, and which is in decided contrast to the tall, more ‘‘racemose’’ (al- though these may be ‘‘ecorymbose’’ at the top) forms such as alba and virginica.

In height angustifolia varies from 75 to 120 em. The central axis develops its corymbose panicle of short racemes first, but it is usually soon overtopped by the successive laterals developed basipetally, each

464 Unversity of California Publications in Botany. [ VoL. 5

lateral, in turn, developing a corymbose cluster of racemes, rising more or less above its predecessors. The result is that the whole plant has the short corymbose habit mentioned above. The stems and branches of angustifolia are comparatively slender, being much more slender than those of macrophylla, or those of any other of the Tabacum varieties except those of calycina, which are very similar.

The leaves of angustifolia are alternate and distinctly and moder- ately long. petiolate. The blade of the lower leaves is ovate-lanceolate,; tapering above to a long, eurved point, more or less conduplicate below and with the rounded bases unequal. Above, the leaves are less con- duplicate, more so even at the base, with the petiole shorter, while the uppermost (bracts) become almost sessile and narrowly lanceolate even to almost linear in outline. The normal petiole is naked at the base and in the middle portion, but the base of the blade is slightly and narrowly decurrent along the upper portion. Occasionally a petiole shows a narrow wing throughout its length and at times the petioles of all the leaves on certain plants are more or less winged, but the majority of the plants have naked petioles (ef. also Goodspeed and Clausen, 1917, p. 306, pl. 46, right-hand figuze). The leaves of angus- tifolia have also a very characteristic drooping habit, much more pro- nounced than in any other Tabacum variety except calycina. In older plants, after capsule formation has well advanced, all the leaves are hanging obliquely downwards.

The flower of angustifolia is distinctive and differs in details of shape and color from that of any other Tabacum variety, and especially from that of macrophylla. The general shape of the flower is that of all the Tabacuwm section, but the corolla is much more slender and more gradually infundibuliform than that of any of the other varieties re- ported here. The calyx is broadly campanulate, prolonged above into 5 long, but unequal, linear-lanceolate, pointed lobes, of which one is longer than the remaining four and gives the calyx a zygomorphie appearance. The corolla is narrow and tubular below the middle, expanding rather gradually and evenly above into a conical infundi- bulum which bears the spreading, deeply 5-lobed limb at its summit. The length of the tube of the corolla is about 6 em. and its greatest diameter about 7 mm. The limb of the corolla, at first erect (opening bud), then horizontal, finally becomes somewhat deflexed and measures about 3 or 3.5 em. aeross. It is divided almost to the tube into 5 lobes which are ovate-lanceolate with long, narrow, tapering tips. The lobes of angustifolia are much longer and have narrower tapering tips than

1922 | Setchell-Goodspeed-Clausen: Nicotiana Tabacum 465

those of any other Tabacum variety, and in this respect are in direct contrast to those of macrophylla. The lobes are unequal and give a slight suggestion of zygomorphism to the corolla. The stamens are inserted on the lower portion of the ‘tube and are usually slightly ex- serted in anthesis. The pistil possesses the usual 2-celled ovary, long, slender style, and thick, slightly bilobed stigma, more or less exserted in late anthesis, characteristic of the genus Nicotiana. The color of the corolla is a light, tlrough lively, pink, much lighter than-the red of macrophylla. The capsule at maturity is slightly flattened longitud- inally, is broadly lanceolate in profile, tapers above into an acuminate apex, and is about 25 mm. high and 8 to 9 mm. thick. It is the most slender of all the capsules borne by the various Tabacum varieties and in decided contrast to the stout eapsule of macroplrylla.

In plate 55, figure -1, is illustrated a plant of angustifolia at the height of its blooming period, Typical features of the plant are shown in the line drawings of plate 56. Photographs of typical leaves are shown in plate 58, where they may be compared with photographs of the leaves of macrophylla. Photographs of the flowers are reproduced in plate 60, where they may be compared with those of macrophylla and of the hybrids between these two varieties.

Macrophylla, U. b. B. G. 22/07, has ie been discussed and figured by one of us (ef. Setchell, loc. cit.). The original seed was obtained from Comes, but the plants do not correspond to his figures (cf. Comes, 1899, pl. VIII) either as to habit or shape of leaf. They differ also from his description in these same respects. The flower, however, agrees, and it seems best to retain for it the name under which we have cultivated it.

The habit and height of macrophylla are both very similar to those of angustifolia. The habit is low corymbose, the central axis bearing a panicle of corymbose racemes and the laterals arising one after the other bearing similar inflorescences and equaling or overtopping the central axis. The stems and branchits are stouter than those of angus- tifolia, however, and this, together with the broader, more solid looking leaves which do not droop so much as those of angustifolia, give a mature plant of macrophylla a much more robust appearance than is the case with a mature plant of angustifolia. The plant figured in the first number of this volume (pl. 6) was young. An older plant shown herewith on plate 55, figure 2, is in full blossom and beginning to ripen its capsules, and gives a better idea of the habit of a well grown plant.

466 University of California Publications in Botany [ Vou. 5

The leaves of macrophylla are sessile by a partially clasping base and possess two basal lobes partially clasping the stem. The general shape is obovate, the widest portion being above the middle. The leaves taper gradually to the broad elasping base below and abruptly to a narrow more or less acuminate tip above. The surfaces show the secondary veins branching at a more obtuse angle than do those of the leaves of angustifolia. The color of the leaves is a dark green in macrophylla and more of a yellowish green in angustifolia. In every way, then, the leaves of the two parents differ from each other as much, in fact, as do the leaves of many species.

The flowers of macrophylla, while of the same general type as those of angustifolia, differ in details of shape and color. The flowers of macrophylla are about 4 em. long. The calyx is broadly ovate in pro- file, deeply cut into 5 broad and somewhat unequal lobes. The corolla tube is stout cylindrical (about 5 mm. in diameter) below, broaden- ing suddenly into a stout infundibulum above (about 10 mm. in diam- eter). The limb is at right angles to the tube, is about 23 mm. across, and is more or less pentagonal with 5 shallow sinuses. The color of the corolla is deep red fading to an almost lilae tint after anthesis. On the limb are 5 triangular lighter areas, one having the narrow apex at each sinus and the broad base at the top of the tube. In the much darker color, in the broader tube and stouter infundibulum, and in the barely appreciable lobing of the limb, the corolla of macrophylla is the oo that of angustifolia. In stamens and pistil, the flower of macrophylla shows little variation from that of angustifolia. y to

The capsule of macrophylla is broadly ovate, tapering abr i ter,

a mucronate tip. It is about 2 em. high and about 1.5 em. in contrasting very decidedly with the comparatively slender capsule

angustifolia.

A typical plant of macrophylla is shown in plate 55, figure 2. Typical features of the plant are shown in line drawings in plate 57. Photographs of leaves are reproduced in plate 58, where they may be compared directly with those of angustifolia. In plate 60 its flowers may be compared directly with those of angustifolia and with those of the hybrids.

It has seemed best to call attention to the characters of and differ- ences between these two varieties, parents in the first set of crosses to be discussed, in order that the behavior of their hybrid progeny may be clear. In height and habit there is a close agreement, but in leaf, flower, and fruit there are sufficient differences to mark them as sep- arate species.

1922] Setchell-Goodspeed-Clausen: Nicotiana Tabacum 467

2. F, OF THE ANGUSTIFOLIA-MACROPHYLLA SERIES

In late July of 1909, some 7 crosses were made between angusti- folia and macrophylla, 6 of which, as stated above, were successful. H,, H,, and H,, as they were designated, involved angustifolia as the female plant, while H,, H,, H,,, and H,, were reciprocals. No seed was obtained from H,, but all the other 6 crosses gave a fair yield. The usual care (ef. Goodspeed, loc. cit., pp. 129-131) was taken in cleaning and sowing the seed. This was done in the spring of 1910, germination was good in all eases, and 337 plants, distributed as follows, came to maturity and seed bearing. The family of H, had 56, H, had 60, H, had 47, H, had 58, H,, had 55, and H,, had 61 plants.

A survey of all these plants showed in general a remarkable uni- formity in habit. A certain amount of difference was to be detected on careful scrutiny, but little if any greater than that which is ex- hibited among a large number of individuals of one or the other parent. In height, F, showed exactly the same variation as the parent, the central axes varying from 65 to 145 em., but largely varying from 90 to 120 em., while the laterals rose to 150 em. Some rows showed uni- formly higher, others uniformly lower plants, the differences probably being due to different soil and water conditions. The habit (see pl. 61) was low corymbose and the general appearange as to stoutness seemed more or less intermediate, between the two parents. The leaves in ‘shape, size, etc., very closely resembled those of angustifolia. There was some appreciable variation in the leaves, however, aud often con- siderable variation on the same plant, a characteristic of angustifolia which has already been mentioned. Plate 59 reproduces photographs of different types of leaves obtained from F, plants. The blade is

broadly elliptical ovate with the lateral veins at an obtuse angle, much

as in macrophylla. The base is rounded, or even slightly cordate in some leaves, while the tip is more blunt. These characters seem, at least, to indicate an influence of macrophylla. The leaf, however, is distinetly petiolate, but the petiole is not so long as in angustifolia.

The petiole is definitely winged and the wings are expanded at the base into auricles, which are often triangularly decurrent along the internode of the stem. This wing is usually present in all the plants of F,, but some leaf or leaves on a plant may lack it, and in some plants it is only slightly developed, or at least, is without auricles. The wing was from 5 to 7mm. wide on some leaves.

The leaf of 10F,H,,P, represented in plate 62 even more closely resembles the typical leaf of angustifolia. The wing along the mar-

468 University of California Publications in Botany [| VoL. 5

gins (or edges) of the petiole is narrow and is prolonged as a slight ridge to the internode and is decurrent (?) or ean be traced as it bends sharply downwards. Such wings are, at times, found in pure bred angustifolia plants. This leaf, then, resembles the angustifolia leaf fairly closely, but differs from its ordinary expression in the more tapering base, in being less distinctly conduplicate, in tapering more abruptly toward the tip, in having shorter petioles and in having more of a wing on the margins of the petiole.

The flower of F, (see pl. 60) resembles that of angustifolia more than that of macrophylla. The color is deep pink, decidedly of a deeper shade than is the flower of angustifolia, yet far from the red of macrophylla and in a way intermediate between the two. There is no trace, on the limb of the corolla of F,, of the 5 white triangle- shaped areas so characteristic of the limb of the corolla of macrophylla. The infundibulum, while possibly slightly stouter than that of the flower of angustifolia, is not so stout as that of the flower of macro- phylla. In length the flower of F, averages about 4 to 4.5 em. as against an average of 6 em. in angustifolia, and of 4 em. in macro- phylla. The tube averages about 3.5 to 5 mm. in diameter below, as contrasted with 2.5 to 3 mm. as an average in angustifolia and 5 mm. in macrophylla. ¥The infundibulum in the corolla of F,, while neither abrupt nor so i," at of the flower of macrophylla, is noticeably more abruptly enlarge@ and stouter than that of angustifolia. The limb of the @orolla in F, averages 2.5 to 3.25 em. in greatest diamete while that

are acta broad at the base, particularly so as compared with their length. In general, then, the corolla of F,, while closer to that of angustifolia, shows by its stouter tube, more abrupt and more swollen infundibulum, intermediate spread of limb, less deep lobing, shorter and broader lobes, and deeper shade of pink, definite influences of macrophylla also. ;

The capsule of F, is broader than that of angustifolia, but nar- rower than that of macrophylla. There is greater variability in hori- zontal diameter in the capsule of F,. The flower and fruit of F,, then, although no careful biometric study has been made, are inter- mediate between those of the two parents, yet incline more toward angustifolia than toward macrophylla.

1922] Setchell-Goodspeed-Clausen: Nicotiana Tabacum 469

In general, then, a survey of F', shows throughout a series of ten different families a uniformity of individuals as great as that exhib- ited in either of the parents. Some few slight differences exist among individuals both of F', and of the parents which may possibly be re- ferred to lack of a completely homozygous conditon in the parents. In characters in which the two parents differ, whether in color of flower, quantitative corolla character complexes, capsule character com- plexes, or leaf character complexes, the F, hybrid exhibited through- out a character expression intermediate between that of the two parents.

3. F, OF THE ANGUSTIFOLIA-MACROPHYLLA SERIES

In 1911, there were selected as parents for the F, 13 plants from H, and 12 from H,. Twenty-one families of approximately 50 plants each were set out in the field. On account of the great diversity shown in these populations, it was found impossible to study individually each of the thousand plants grown; consequently particular attention was paid to only 5 families from each hybrid. The other families were gone over carefully, but nothing notably different was found in their behavior. All fifty plants survived in each family except in the last,

, 11F,H,P,,, where only 48 came to maturity. There were then is plants of F, under more careful observation, representing both the eross and its ‘reciprotille with about 550 remaining for only easual examination.

As might have pat expected, there was 4 ereat variety of plants resulting and segregation as to differences in combination of characters of flower, fruit, and leaf was little short of bewildering. An attempt was made to study and jarrange these combinations, but it was found to be impossible. A eareful survey, however, was nade of the popu- lations and a sha arate was attempted. Wome 16 fairly réadily separable types, based on leaf characters, were distinguished, but between these closely approaching types others were to be found of intermediate and overlapping character. One each of the types selected was drawn, and these drawings are reproduced in plates 63 to 78.

A glance at these plates, which were carefully drawn to scale, will show something of the nature of the combinations of characters of the two original parents. Type 1 (pl. 63) shows a close approximation, yet not an absolute reproduction, of angustifolia, while type 16 (pl. 78) in a similar way is a close approximation to macrophylla. The other

470 University of California Publications in Botany [ Vou. 5

14 types (pls. 64 to 77) are clearly intermediates approaching one parent more than the other, but types 12, 13, and 14 (pls. 74 to 77, inclusive) are decidedly different from either as to leaf, at least, and type 10 (pl. 72) is of another altogether different form, although all of these leaf shapes are connected to a greater or less extent into one series of more or less gently intergrading forms.

As to the shape and dimensions of the corolla there is to be found a similar series of intergrading forms from the slender corolla tube with gradually expanding and slightly swollen infundibulum and deeply lobed limb of type 1 (pl. 63) to the corolla with stout tube, abruptly and considerably swollen infundibulum with slightly lobed limb of type 16 (pl. 78). In color the corollas vary from the light pink of angustifolia to the red of macrophylla and three shades are at times fairly readily distinguishable, the light pink of angustifolia, the deep pink of F',, and the several nuances of the red of macrophylla.

The capsules also show various combinations from the slender gradually attenuated capsules of angustifolia to the stout, swollen, abruptly upwardly attenuated capsules of macrophylla. Both ecap- sules and corollas approaching one parent may be found with leaves more closely approaching the other parent. In stature and habit the plants of all the | milies were reasonably uniform and agreed in general in these respects*with the parents and F,, there certainly being no greater amplitude of?variation in _ than was to be found in the parental types. ' ’

Among the great variations, two charactérs _seemed to stand out fairly \clearly for, a statistical = viz., color of the

?

corolla ‘nd the possession, or lack, of a petiole. Numerical data: for these characters are given in table 1. Some ¢are was taken to obtain ~~ © as regards ea¢h of the characters. As was, as noted before, possible to distinguish shades, or sets of shades, which were designated as light pink, pink,

a careful

regards color

red. In practice, however, it was usually difficult to distinguish * two shades of pink from each other. The red gave very little trou

In attempting to classify the plants of F,, with respect to type of leaf base, more difficulty was experienced because of the variety of forms which were produced and the degree of intergradation which existed between forms. In judging the presence or absence of petiole, therefore, in these populations, the classification is faulty because of lack of knowledge of the genetic constitution of the various distinct forms and those which grade into them. In table 1 the plants are

~

*

I

1922 | Setchell-Goodspeed-Clausen: Nicotiana Tabacum 471

thrown into the petiolate class if they were distinctly narrowed at the base, and whether naked or winged.

In F,, then, there appears to be simple Mendelian inheritance in only one pair of the original character contrasts of the parents, namely, red versus pink corolla color. Here the hybrid is intermediate and F’, segregates sharply into pink and red in the ratio 3 pink : 1 red. Within the pink class there is a more or less evident segregation into 2 pink : 1 light pink, but the shades intergrade so that no distinet line of demarcation exists between the classes. As respects leaf base char- acters, the segregation is so complex that no reasonable genetic analysis is possible. The numerical data for this latter character presented in table 1 are of value only in that they indicate a close agreement in segregation among F’, families, thereby furnishing a rough statistical demonstration of the equivalence of the several familes. The more definite data on leaf base characters are derived from generations subsequent to F,. %

TABLE 1 v

CLASSIFICATION OF F2 PLANTS OF THE ANGUSTIFOLIA-MACROPHYLLA SERIES ACCORDING TO COROLLA COLOR AND LEAF BASE G@HARACTERS.

= a

Corolla Color r Leaf Base Family Garden F CEU) eeonbers f d pink light pink petiolate non-petiolate A 11F,H2P. 12 23 14 42 7 B 11F.H2P; 14 25 10 30 20 Cc 11F,H.Ps 13 21 15 34 14 D 11F.H»2P, a 21 22 35 15 E 11F,H>Pi; 15 20 14 30 20 F 11F.H,P, 13 28 a 32 17 G 11F,H,P;; 12 29 8 35 ) ait 14 H 11FHyPy 13 1 22 32 ae 18 J LRELPs: 8 bear | io 31 17 Kk 11 FH yPas 6 27 15 38 10 Totals 113 240 137 339 » BY

472 Umiversity of California Publications in Botany [ Von. 5

4, F; AND SUBSEQUENT GENERATIONS OF THE ANGUSTIFOLIA-— MACROPHYLLA SERIES

From F, of H, and H,, 20 plants were selected for further ex- perimentation and families of 25 were determined upon as the unit. In all except four, the families of 25 each were successfully raised. Of one of the four only 14 plants were obtained, which were all that germinated, while in each of the other three families 24 plants were reared to maturity. Altogether, then, 486 plants were raised of the F, during the season of 1912. It was the intention to grow from each of the selected types, as drawn for illustration. Fifteen of the families from the type parents were successfully reared, but, in some way or other, the seed of type 4 (10F,H,P,P,,) was not to be found, and, unfortunately, no note had been made as to whether or not any seed was produced. No complete sterility, however, was noticed in any members of F, of either H, or H,, and the presumption is that F', seed of type 4 must have been lost in har vesting.

The variation within each family was decidedly less than that of the families of F,. Of the 20 families reared wholly or in part, 4 families were rete uniform, varying in minor details only. Five families segrega only in coroll lor, 4 segregated only in leaf base characters, and “fle remaining 7 segregated both in ecoroll color and leaf base characters. In table 2 are summarized the de as