Genetic studies in populations of parviflora Dougl.ex Lindl. ()

by

Gerda Rosa Krause Sc. (Hon.), University of British Columbia, 1975

A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science

in The Faculty of Graduate Studies Department of

We accept this thesis as conforming to the required standard

The University of British Columbia

Gerda Rosa Krause, 1978 In presenting this thesis in partial fulfilment of the requirements for an advanced degree at the University of British Columbia, I agree that the Library shall make it freely available for reference and study.

I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the Head of my Department or by his representatives. It is understood that copying or publication of this thesis for financial gain shall not be allowed without my written permission.

Department of

The University of British Columbia 2075 Wesbrook Place Vancouver, Canada V6T 1W5

Date - ii -

Abstract

The Collinsia populations in southwestern British Columbia and northwestern Washington show considerable variation in leaf and flower characters both within and between populations. One of the purposes of this study was to determine the genetic mechanisms con• trolling some of these characters.

Two leaf polymorphisms were studied. They were the presence or absence of purple anthocyanin spots on the surface of the leaves and the presence or absence of a silvery sheen, also on the surface of the leaves. Each was shown to be controlled by a single gene with two alleles. The spotted leaf character was dominant over the un• spotted leaf character and the silvery sheen was dominant over the normal green leaf character.

Flower size was also studied and shown to be controlled by polygenic inheritance.

Two mutant flower colours, white and magenta, are found in this region of study, in addition to the normal blue colour. The inher• itance of flower colour was not conclusively determined but the data indicate that two genes may be involved, one controlling the pro• duction of the magenta pigment from the colourless precursor and one controlling the production of the blue pigment from the magenta one.

Another purpose of this work wa6 to determine the chromosome number of the Collinsias in this area. Both diploid (n=7) and tetraploid (n=l*f) counts have been reported. Six populations were studied and all were found to be tetraploid. - iii -

Finally, the cytological and genetic data were used in con• junction with morphological data to revise the of the

Collinsias in this region of study. Most authors divide them into two , £. grandiflora Dougl, ex Lindl, and parviflora Dougl, ex Lindl, However this study indicates that they are only one highly variable species, C_, parvi flora. - iv -

Table of Contents Page

Abstract ii Table of Contents iv List of Tables v List of Figures and Illustrations vii Acknowledgement ix

Chapter 1 Introduction a) Taxonomy 1 b) Cytogenetics 2 c) Genetics 3 d) Materials and Methods 5

Chapter 2 Leaf Spot Polymorphism a) Introduction 10 b) Materials and Methods 13 c) Results 15 d) Discussion 21

Chapter 3 Silvery Leaf Polymorphism a) Introduction 24 b) Materials and Methods 2If c) Results 26 d) Discussion 33

Chapter k Flower Colour Mutants a) Introduction 3k fc) Material and Methods 38 c) Results ifO d) Discussion k9

Chapter 5 Inheritance of Flower Size a) Introduction 51 b) Materials and Methods 53 c) Results 57 d) Discussion 73

Chapter 6 Chromosome Counts a) Introduction 77 fc) Materials and Methods 78 c) Results 78 d) Discussion 79

Chapter 7 Taxonomy a) Introduction 81 b) Materials and Methods 81 c) Results 85 d) Discussion 90

Bibliography Vita - v -

List of Tables

Page

Table I Population numbers of the Collinsia populations used in the crossing experiments •••••••••••••••• 7

Table II Si and F^ progeny of the crosses involving the spotted leaf character • • • 16

Table III S2 and Fp progeny of heterozygous spotted S^ and F^ ••••••••• 18

Table IV Sg and Fg progeny of unspotted S^ and Fx plants ••••••• 19

Table V Sg and Fg progeny of homozygous spotted S, and F, plants ••••••••••••• 22

Table VI S^ and F]_ progeny of the crosses involving the silvery-leaf character • • • 27

Table VII S2 and F2 progeny of the crosses involving the silvery-leaf character • • • 29

Table VIII F2 progeny of the #11 green (9) x #25 silvery (d1) cross •••••••

Table IX S^ and So progeny of the populations used in the crosses involving flower colour 41

Table X F^ and Fp progeny resulting from crosses between blue-flowered and white-flowered plants •••••••••••••••••• 43

Table XI F2 progeny of the #22 (?) x #9 blue (<3) cross that showed segregation for flower colour •••••••••••••••••• kk

Table XII F^ and F2 progeny of the crosses between blue-flowered plants and magenta-flowered plants •••••«•••••••••••• 46

Table XIII Fi and F2 progeny of the crosses between white-flowered magenta-flowered plants • • 48

Table XIV Corolla length classes assigned to C, parviflora and C. grandiflora by various authors •••••••••••••• 32

Table XV Populations of C. barviflora used in the study of flower size variation . . • , 55 - vi

Table XVI Mean flower sizes and sample sizes of the populations graphed in Fig. 17

Table XVII Mean flower sizes and sample sizes of the populations graphed in Fig. 18

Table XVIII Mean flower sizes and sample sizes of the populations graphed in Fig. 19

Table XIX Mean flower sizes and sample sizes of the populations graphed in Fig. 20

Table XX Mean flower sizes and sample sizes of the populations graphed in Fig. 21

Table XXI Mean flower sizes and sample sizes of the populations graphed in Fig. 22

Table XXII Mean flower sizes and sample sizes of the populations graphed in Fig. 23

Table XXIII Mean flower sizes and sample sizes of the populations graphed in Fig. 24

Table XXIV Data sheet for population ______

Table XXV Summary of the variation among the populations studied .... • ..• - vi j -

List of Figures and Illustrations

Page

Figure 1 Map of the study area showing the local• ities from which seeds were collected • • • 6

Figure 2 Collinsias growing in 5-inch pots • • • • • 8

Figure 3 Collinsias growing closely spaced in shallow flats ••••••••••••••• 8

Figure k Plants with and "without purple antho- cyanin spots on the upper epidermis of the leaves •••••••••••••••• 11

Figure 5 showing the faint-spotting character ••••••••••••••••• 12

Figure 6 Plant showing the heavy-spotting character ••••••••••••••••• Ik

Figure 7 Carlos Island plants showing the silvery-leaf character •••••••••• 25

Figure 8 Carlos Island plants showing normal

green leaves ••••••••••••••• 25

Figure 9 Silvery, green and intermediate' plants • 31

Figure 10 A magenta flower from the Elk Falls population •••••••••••••••• 35 Figure 11 An albino flower from the Mt» Douglas Park population •••••••••••••• 35

Figure 12 A population with anthocyanin pigments in the leaves ••••••••••••••• 37

Figure 13 The Mt, Douglas Park population with yellow leaves ••••••••••••••• 37

Figure Ik The F2 generation in the cross between white and magenta-flowered plants, showing segregation for the three flower colours • k?

Figure 15 The variation in flower size between the five populations used in study. From left to right: Elk Falls, Jack Point, Nanoose Hill, Botanie Valley, Lindeman Lake • • • • 3k

Figure 16 A Colljnsia flower showing the angle at which measurements were taken • • • • • 56 - viii - Page

Figure 1? Graph comparing flower sizes of the five populations in the first (parental) generation ••••••••••••••••• 58

Figure 18 Graph comparing flower sizes of the five populations in the second (S,) generation • 60 Figure 19 Graph comparing flower sizes of the five populations in the third (S^) generation • • 62

Figure 20 Graph comparing the flower sizes of Lindeman Lake (#6) and Jack Point (#11) populations with a possible hybrid between them •••••••••••••••• 65

Figure 21 Graph comparing the flower sizes of the hybrids between populations from Botanie Valley (#22) and Nanoose Hill (#9) with their parental populations ••••••••• 67

Figure 22 Graph comparing the flower sizes of the hybrids between populations from Botanie Valley (#22) and Jack Point (#11) with their parental populations ••••••••• 69

Figure 23 Graph comparing the flower sizes of hybrids between populations from Lindeman Lake (#22) and Elk Falls (#17) with their parental populations •••••••••••• 71

Figure 24 Graph comparing the flower sizes of hybrids between populations from Mt. Douglas

Park (#2) and Elk Falls (#17) with their parental populations •••••••••••• 74

Figure 25 A pollen mother cell from the Nanoose Hill population showing a chromosome number of 2n s 14 II 80

Figure 26 A pollen mother cell from the Carlos Island population showing a chromosome number of 2n = 14 II 80

Figure 27 The leaf characters of the Botanie Valley population ••••••••••••• 87

Figure 28 The leaf characters of the Jack Point population ••••••••••••••••• 87 Acknowledgement

I would like to express my gratitude to my director,

Dr. F. R. Ganders and to my committee, Dr. C. J. Marchant and

Dr. A, J. F. Griffiths for their willing assistance, discussion and criticism during the research and their help in preparing the manuscript; to Dr. K. I. Beamish, Dr. C. 0. Person and

Dr, B. A. Bohm for advice and assistance with various parts of this thesis; to Mr. Ken Carey and Dr. W. B. Schofield for collecting

Collinsia seeds; and to the Department of Botany for the use of equipment and facilities. I would also like to acknowledge receipt of two National Research Council Scholarships (1975/76 and 1976/77) and thank Mrs. Lorraine Wiebe for typing the manuscript. Finally,

I would like to thank my husband, Eric, for all his encouragement and infinite patience. Chapter 1 Introduction

Taxonomy

The blue-flowered Collinsias in southwestern British Columbia and northwestern Washington have generally been divided into two species. The larger-flowered plants are usually considered C, grand- iflora Dougl, ex Lindl, and the smaller-flowered plants are placed in

C, parviflora Dougl, ex Lindl, (Abrams, 1951; Hitchcock et al,, 1959;

Taylor and MacBryde, 1977)* C, grandiflora was originally described by Lindley (1827) from garden specimens grown from seeds collected by David Douglas in the vicinity of the Columbia River, C^ parvi• flora was described in the same year, again by Lindley, from garden specimens grown from seeds collected by Douglas at "the dry banks of the Columbia River, at the distance of an hundred miles and more from the ocean",

Newsom (1929), in her monograph of the maintained the two species and further recognized two varieties of C_, grandiflora;

C, grandiflora var, typica and C± grandiflora var, pusilla Gray, variety pusilla having smaller flowers than typica. Variety pusilla has also been given subspecific rank (Piper, 1906), Peck (1961) followed Howell (1903) and recognized pusilla at the specific level:

£_t Pusilla (Gray) Howell, Hitchcock et al. (1959) and Pennell

(in Abrams, 195D considered pusilla to be a synonym of C, parviflora« although Pennell recognized two varieties of parviflora. All of the recent floras of the west coast of North America have recognized two species in this group, although they do not always agree on how to separate these entities. There has been little uniformity among authors over where to draw the line between Cj, grandiflora and C, parviflora, This con• fusion is reinforced by the condition in nature. One finds a con• tinuous series of intergrading populations from the smallest flowered parviflora to the largest flowered grand!flora. This fact was noted by Newsom (1929) but she maintained the two species on the basis that

"the, two extremes differ greatly'1. However, after hybridization and morphological studies, I have come to the conclusion that this

Collinsia group containing grandiflora. parviflora and pusilla is only one highly variable species. The name C, parvlflora has priority over the name £• grandiflora. Therefore I will use the name

C, parviflora to refer to the whole group unless otherwise specified.

Cytogenetics

Garber and his students have published a considerable amount of work on the genus Collinsia, Most of the work is cytogenetic, some' is morphological and some is chemical. However C± parviflora« which he divides into parvlflora and grand!flora, has not been examined except

for basic chromosome counts. He reported a chromosome number of n=7

for both species (Garber, 1956, 1958b), However, Taylor and Mulligan

(1968) reported that £• parviflora in the Queen Charlotte Islands had a chromosome number of n=14. The chromosome counts obtained from the

populations in this study agree with Taylor and Mulligan rather than

Garber, Unfortunately, Garber does not give the localities of his

samples nor does he cite voucher specimens. - 3 -

Genetics

Newsom (1929) divided the genus into two groups based on the length of the pedicels. One group of species has sessile flowers, congested into whorls, the other group has pediceled flowers that are solitary or in whorls, Garber (1958a) divided the species into two groups according to whether they exhibited high or low chiasmata frequencies, and these groups correlated well with the*pediceled flower group and the sessile flower group respectively. He also dis• tinguished between "variable" and "uniform" populations (Garber, 1974),

Although he does not define precisely what he means by these terms in his discussion of sessile-flowered, he states that "unpublished data indicate that these (sessile-flowered) species share a large number of phenotypes, each controlled by a single gene difference, not yet observed,,, in the species with distinctly pediceled flowers"

(Garber, 1958b).

On the basis of the above three characteristics, Garber places

C. grandiflora and C_» parviflora into his Group II which contains plants with pediceled flowers, uniform populations and high chias• mata frequency. The collinsias in southwestern British Columbia and northwestern Washington are definitely pediceled. There are no data on the chiasmata frequency. However, the populations are not very uniform. Many populations show considerable variation in leaf size, shape, pubescence and colouring, flower size and colouring, etc. No doubt many of these characters are genetically controlled. I investigated a number of these polymorphisms to determine the genetic mechanisms controlling them. - k -

One common polymorphism is the presence or absence of dark

purple spots on the upper surface of the leaves. This character was found to actually include two separate characters. A gene system

producing large, heavy spots sometimes covering most of the leaf was

found to be a simple dominant. The other character appeared as a few

tiny dots on one or a few leaves of the plant. Its genetic mechanism was not positively determined but the results of a study by Griffiths

et al. (1977) suggests that a single incompletely penetrant gene is involved.

Another leaf polymorphism is the presence or absence of a silvery

sheen on the early leaves. This was also shown to be controlled by a

single gene with a dominant allele for the silvery sheen.

The inheritance of flower colour is more complex. Three different

flower colours were found in the parviflora populations studied.

These were magenta and white in addition to the normal blue. The results were.not conclusive but suggested the following genetic

interpretation. One gene (A) controlled the production of the magenta pigment from the colourless precursor. Another gene (B) controlled

the production of the blue pigment from the magenta pigment. A homo•

zygous recessive (aa) at the first step resulted in a white-flowered plant and a homozygous recessive (bb) at the second step resulted in a magenta-flowered plant. This relatively simple Mendelian system

waB, however, complicated by selection against the recessive alleles

in the gamete or zygote which distorted the expected 3*1 ratios.

The genetic control of flower size is of special interest

because Cj, parviflora and C. grandiflora are separated mainly on the - 5 -

basis of that character. As expected for a continuously varying character such as flower size the inheritance mechanism is probably polygenic. However it was not possible to determine exactly how many genes are involved.

Materials and Methods

C. parviflora is a good organism for genetic studies since it grows well under artificial conditions and, being an annual, has a relatively short life cycle.

The populations of plants used in this study were grown from seed

collected from various places in southwestern British Columbia and northwestern Washington. Figure 1 is a map of the area showing the

localities from which seeds were collected. For convenience, the

populations used in the crossing experiments were assigned numbers.

These population numbers are listed in Table I.

Seeds were collected from mature brown capsules and stored in

paper packets until needed. Seeds were sown between pieces of moist

filter paper in petri plates and placed in a refrigerator at 5° - 9°C

until the seeds germinated and the radicles were about 1-2 cm long.

This usually took about two to three weeks. They were then planted

in flats or pots of soil. The parental and F^ generations were grown

in 5 inch pots (Fig. 2) and were large vigourous plants, but since

the number of individuals in the F2 generation was usually much larger,

it was planted closely spaced in shallow flats (Fig. 3)« As a result,

the plants were often smaller and less vigourous but this did not

affect the study because size and vigour were not measured and the - 6 - Table I

Population numbers of the Collinsia populations used in the crossing experiments

Source of seeds Population number (Locality)

#2 Mt« Douglas Park, Vancouver Island

#6 Lindeman Lake, B. C,

#9 Nanoose Hill, Vancouver Island

#11 Jack Point, Vancouver Island

#17 Elk Falls, Vancouver Island

#22 Botanie Valley, B, C.

#25 Carlos Island, B. C. - 8 -

Fig. 3 Collinsias growing closely spaced in shallow flats - 9 - characters studied were expressed in the small plants as well as the large ones. Three to four weeks were required to grow seedlings in soil to the rosette stage where leaf polymorphisms could be evaluated and another four to five weeks before most of the plants started to flower.

All plants were grown in growth chambers under a cool, long day regime of 16 hours light at 20°C and 8 hours dark at 10°C. This regime proved best for good vegetative growth and flowering and the low temperature ensured maximum expression of the spotting character.

Controlled cross-pollination is quite difficult because the flowers are small and will self-pollinate readily. They have to be emasculated in the bud stage before the pollen is mature and before the anthers can dehisce either naturally or accidentally with handling.

The buds are only about 2-3 mm long, and must be handled under the dissecting microscope in order to ensure that the anthers are com• pletely removed and the style and stigma are not damaged. An additional complication is that occasionally one of the anthers will dehisce in the closed bud stage and scatter pollen over the stigma.

However, in a successfully emasculated bud, the stigma will reach maturity in 3 to 5 days and can then be cross-pollinated. This was accomplished by removing mature anthers from a flower of the male parent plant and brushing them over the receptive surface of the stigma. - 10 -

Chapter 2 Leaf Spot Polymorphism

Introduction

One of the most obvious polymorphisms found within many of the populations of on Vancouver Island is the pres• ence or absence of purple anthocyanin spots on the upper epidermis of the leaves (Fig. 4). These spots vary in size, number and shape both among and within the individual plants and are usually found only on the earlier leaves of the plant and sometimes on the cotyledons. The anthocyanin spots usually fade and disappear as the plant gets older or as the temperature rises. For example, wild collected spotted plants transferred to a heated greenhouse lost their spots within three days.

Gorsic (1957) reported a similar polymorphism in his genetic studies of (Lj, heterophylla Buist. He called it dark-dotted (Ld) and described it as a transient cotyledon and leaf character in which the

"upper surfaces of cotyledons and leaves show maroon spots of various numbers and sizes". He found this character to be inherited as a simple dominant.

Griffiths et al., 1977» in working with populations of £. grand- iflora Lindl. in southwestern British Columbia and northwestern

Washington discovered two genetically distinct spotting systems, a faint spotting system (F) and a heavy spotting system (H). There is considerable variation within each system and the ranges of the degree of spotting within the two classes probably overlap. However, plants with the faint degree of spotting usually have only a few of their leaves bearing a small number of tiny dots, about 0.5 mm in diameter

(Fig. 5). The precise genetic determination of the faint spots was not conclusively established but the results suggested that a single - 11 -

Fig, k Plants with and without purple anthocyanin spots on the upper epidermis of the leaves - 12

Fig, 5 Plant showing the faint-spotting character - 13 -

incompletely penetrant gene is involved. Plants with a heavy degree of spotting usually have blotches of various shapes and sizes on all or most of the leaves and on the cotyledons (Fig, 6), The amount of blotching can vary considerably from a large blotch covering almost the entire leaf to a few small spots near the base. The genetic determination of heavy spotting is distinct from that of faint spotting and maintains its inheritance pattern even when the faint-spotting gene is present.

The spotting system investigated in this study was the heavy spotting one. Crosses were set up to determine the exact inheritance pattern for this particular polymorphism.

Materials and Methods

In order to study the genetics of the spotting system, an exper• imental population containing both spotted and unspotted plants was used. Reciprocal crosses were set up between spotted and unspotted plants from a population of plants (population #11) grown from seeds collected from Jack Point (Fig, 1), Spotted and unspotted plants were also allowed to self. In addition, reciprocal crosses were made between the spotted plants of population #11 and plants from another population (population #9), derived from seeds collected from Nanoose

Hill (Fig, 1), that contained only unspotted plants. These unspotted plants were also allowed to self.

In order to get sufficient progeny from the crosses, more than one plant was involved in each cross. Therefore, both homozygous and heterozygous parents could have been involved in any particular cross or self. Fig, 6 Plant showing the heavy-spotting character - 15

Seeds from the cross-pollinations were collected, germinated and grown to the rosette stage when they were classified and tagged for the presence or absence of spotting in the early leaves. The tagging at this stage is very important since the spotting fades as the plant gets older. The plants were then left to self naturally and the seed from each of these plants was collected and grown individually to be scored.

Results

The results of the artificial crosses i.e, the F^ plants and

the results of the self-pollinations, i,e, the S^ plants are

summarized in Table II,

These results indicate that the absence of spots is a true-

breeding character in populations from both Nanoose Hill and Jack

Point, The spotted plants on the other hand, produced both spotted

and unspotted progeny when selfed, indicating that a single gene

system may be involved and that at least some of the plants involved were heterozygous for spotting and that the spotted characteristic

is dominant over the unspotted one.

The indication that the spotted character is dominant is rein•

forced by the reciprocal crosses. When the spotted plants are used

either as male or as female parents and crossed with unspotted plants

the resulting progeny are mostly spotted. All of the spotted F-^

progeny would be expected to be heterozygous, containing spotted

alleles from their spotted parents and unspotted alleles from their

unspotted parents. Table II

S.. and F, progeny of the crosses involving leaf spots

Pr Pollination regime e °Seay ° Spotted Unspotted

Population #11; spotted selfed 18 5

Population #11; unspotted selfed 0 22

Population #9; unspotted selfed 0 13

#11 spotted (9) x #11 unspotted (cf) 26 15

#11 unspotted (?) x #11 spotted (o") 22 7

#11 spotted (9) x #9 unspotted (cf) 13 11

#9 unspotted (?) x #11 spotted (cr) 15 19 - 17 -

Seeds were collected from each of the and S^ plants that reached maturity (some were killed by aphids and mildew) and each family was grown and scored individually to determine the genetic make-up of the parent and to determine the ratio of spotted plants to unspotted plants in the progeny of the heterozygotes.

Most of families grown from spotted F^ and S^ plants segregated into spotted and unspotted plants in approximate y.l ratios confirming that they were heterozygotes and that a single gene with a dominant allele for the presence of heavy spotting and a recessive allele for the absence of heavy spotting controls this particular polymorphism.

The actual ratios are summarized in Table III.

Most of.the heterozygotes seem to fit the 3:1 ratio well and confirm that heavy spotting is inherited as a simple dominant. The

F2 progeny resulting from the #11 spotted (9) x #9 unspotted (cf) cross, however, shows a slightly significant deviation from the expected ratio due to a greater number of unspotted progeny than expected. This is typical of the behaviour of the faint-spotting allele (Griffiths et al., 1977) and can probably be attributed to one or more faint-spotted plants having been mistaken for a heavy- spotted plant.

Table IV summarizes the F2 and progeny of all of the un• spotted plants resulting from both cross and self-fertilization.

If unspotted plants are really homozygous recessives all of the progeny of unspotted plants would be expected to be unspotted. This is not entirely the case. Although the vast majority of the progeny are unspotted, there are a few anomalies. However, since these anomalies are few and do not seem to fit any particular ratio, it is Table III

S2 and F2 progeny of heterozygous spotted S1 and F1 plants

# Of S2 & F2 Number of S2 & F2 individuals Origin of spotted and F^ plants families Spotted Unspotted X2 (1 d.f.)

Population #11; spotted self 261 100 1.40; P=0.5-0.1 deviation not significant

#11 spotted (?) x #11 unspotted (d1) 10 619 184 1.87; P=0.5-0.1 deviation not significant

#11 unspotted (?) x #11 spotted (c?) 18 466 132 2.73; P=0.1-0.05 deviation not significant

#11 spotted (?) x #9 unspotted (c?) 12 250 111 6.36; P=0.025-0.01 deviation significant

#9 unspotted (9) x #11 spotted (

S2 and F2 progeny of unspotted and F^ plants

# of S2 & F2 Origin of unspotted # of S2 & F2 Individuals S^ and F^ plants families Spotted Unspotted

Population #11; spotted selfed 3 5 312

Population #11; unspotted selfed 3 o 265

Population #9; unspotted selfed 10 o 404

#11 spotted (9) x #11 unspotted (

#11 unspotted (9) x #11 spotted (c?) 7 40 698

#11 spotted (9) x #9 unspotted (c?) 10 1 685

#9 unspotted (?) x #11 spotted Q?) 7 2 417 20 -

probable that they are not a result of the heavy-spotting genetic system. Some, for example, were probably contaminants resulting when the seeds were planted. The families were sown side by side in plastic"flats and a seed from a spotted family could quite easily have floated over the barrier to an unspotted family when they were being watered. This is probably the case in the cross #11 spotted x #9 unspotted (&) where out of 10 families, 9 showed only unspotted plants and 1 family had a single spotted plant growing close to the barrier between the unspotted family and a spotted one. The same argument can explain most of the other anomalies.

However, one F2 family grown from an F^ plant from the cross,

#11 unspotted (§) x #11 spotted fcT) had 37 spotted and 94 unspotted

F2 plants. This is too large a number of spotted plants to be ex• plained simply as contamination of a strictly unspotted family. But this is also too small a number of spotted plants to be explained as normal 3*1 segregation of a heterozygote spotted plant. The parent plant of this family was originally labelled 'spotted* but because of the high percentage of unspotted plants it was probably not spotted, at least not heavily-spotted. It seems more reasonable to consider this a faint-spotted plant. In general, obviously faint-spotted plants were scored as 'unspotted' but the amount of spotting in this family falls into the range of overlap between the heavy and faint- spotting characters and so was misinterpreted.

If the above explanations for the anomalies are accepted, un• spotted plants can be considered to be homozygous for the recessive character, absence of heavy spots. 21 -

Some of the families scored also showed all spotted and no unspotted progeny indicating that the parent plants were homozygous for the dominant character, heavy-spotting. The results are summarized in Table V.

According to these results, six of the S^ plants resulting from the selfing of spotted plants from population #11 (Jack Point) were homozygotes. This was not unexpected. But, all of the plants resulting from the #11 spotted ($) x #11 unspotted (cf) cross should have been heterozygotes. This apparent anomaly can be explained by noting that the seed parents in the controlled cross were spotted plants. An imperfect emasculation can result in a self-fertilization producing a homozygous spotted plant. Since emasculation of Collinsia parviflora buds is so difficult, these accidental selfs are not unusual. The 6 unspotted plants found in these 2 families are probably the result of seeds floating in from the adjacent unspotted families.

Discussion

It.can be concluded from the above results that the heavy- spotting polymorphism in £. parviflora is controlled by a single

Hendelian gene with a dominant allele for the presence of heavy spotting and a recessive allele for the absence of heavy spotting.

Another.allele, that for faint-spotting, may be involved at this locus (Griffiths et al.) but the actual inheritance pattern of this particular character remains in doubt and requires further research. Table V

S2 and F2 progeny of homozygous spotted and plants

# of S2 & F2 # of S2 and F2 individuals Origin of spotted and F^ plants families Spotted Unspotted

Population.#11; spotted selfed 491 0

#11 spotted (9) x #11 unspotted (cf) 2 93 6 - 23 -

It is interesting to note that the heavy-spotting gene (H) in

C. parviflora shows the same type of inheritance pattern as the dark- dotted gene (Ld) that Gorsic (1957) described in heterophylla.

This similarity in inheritance pattern as well as the similarity in his description of the dark-dotted character makes it appealing to conjecture that the genes involved in both species may be homologous.

However, there is no clear evidence that this is the case.

Now that the genetic determination of the heavy-spotting character is.understood, it can be used to determine outcrossing rates using progeny tests within populations of C_» parviflora.

Griffiths et al. (1977)» in surveying populations of Collinsia in southwestern British Columbia and northwestern Washington found the spotting character to be quite common in the area between Crofton and Little River on Vancouver Island and in those populations in which it does not occur naturally it could easily be introduced since different populations of parviflora are able to interbreed

(see Chapter 7)* Spotted plants can be distinguished in the field with relative ease, especially the strongly heavy-spotted ones, as long as they are scored and marked when in the rosette stage. The

faint-spotting phenotype can be a complication when the degree of

spotting falls into the range of overlap between the two types of

spotting. But since errors in scoring could occur in either direction,

a large enough sample could eliminate any major deviations. Therefore,

the heavy-spotting gene could become a useful tool in determining

breeding systems and outcrossing rates within populations of

Collinsia parviflora. / - 24 -

Chapter 3 Silvery Leaf Polymorphism

Introduction

Another interesting leaf and cotyledon polymorphism was found in a population of CL* parviflora grown from seeds collected on Carlos

Island, B. C. (Fig, 1), In this population some of the plants had a wrinkled upper surface and silvery sheen on their earlier leaves and sometimes on their cotyledons (Fig. 7). Other plants had normal non-wrinkled, green leaves and cotyledons (Fig. 8). In cross-section these green leaves looked normal with a typical, closely-packed palisade cell layer. The silvery leaves, however, had a very loosely packed palisade layer, full of air pockets. When a drop of water was added to a freshly cut silvery leaf cross-section, the epidermal cell layer would float away from the rest of the leaf section as the palisade layer broke up.

Like the spotted-leaf character, the silvery-leaf character is transient and generally seems to fade or be masked by anthocyanin pigment as the plant ages. Unlike the spotted-leaf character, how• ever, the silvery-leaf character is very difficult to distinguish in nature, possibly due to masking by the anthocyanin pigments in the leaves.

Materials and Methods

The procedure used to determine the inheritance pattern for the silvery-leaf polymorphism was very similar to that used for the spotted-leaf polymorphism. Silvery and green plants were chosen from population #25 which was grown from seeds collected on Carlos Fig, 8 Carlos Island plants showing normal green leaves - 26 -

Island and reciprocal crosses were set up between them. Silvery plants from population #25 were also reciprocally crossed with plants from population #11 (Jack Point), which contained no silvery plants (Fig. l). All of the parent plants were also allowed to self.

Controlled crossing using the Carlos Island plants as seed parents was especially difficult. Not only were the buds small and difficult to,work with, the pedicels were so short that the flowers were borne very close to the stem and almost hidden among the leaves.

This made them extremely difficult to manipulate while they were being emasculated. Therefore, more than one plant was used in each cross and self and both homozygotes and heterozygotes may have been involved.

The F^ and S^ plants were grown to the rosette stage, scored and tagged before the silvery character faded. Seed from each of these plants was then grown and scored individually.

Two other plants brought from Carlos Island were crossed in an attempt to study the inheritance of flower colour. The results were inconclusive as far as flower colour was concerned but the F^ and offspring included silvery and green-leafed plants and so were used for the assessment of the silvery-leaf character.

Results

The two plants from Carlos Island used in flower colour crosses showed good silvery-leaf results as mentioned above. The results of the cross-fertilizations and self-fertilizations of the two Carlos

Island plants are summarized in Table VI. Table VI

S, and F, progeny of the crosses involving the silvery-leaf character

# of individuals Pollination regime Silvery Green

Plant A selfed 22 0

Plant B selfed 0 34

Plant A (?) x Plant B (cT) 7 0

Plant B (9) x Plant A (cf) 12 0

Population #25; silvery-leafed, selfed 15 0

Population #25; non-silvery, selfed 0 14

#25 silvery (9) x #25 non-silvery (cT) 5 0

#25 non-silvery (?) x #25 silvery (cf) 0 16

#25 silvery (?) x #11 non-silvery (d1) 5 0

#11 non-silvery ($) x #25 silvery (cf) 15 7 28

These data indicate that a single gene system is probably in• volved and that plant A was a homozygous silvery-leafed plant and that plant B was a homozygous green-leafed plant. The reciprocal crosses indicate that the silvery-leafed character is dominant over the non-silvery-leafed character.

Table VI also summarizes F^ and S^ progeny from population #25

(Carlos Island) and population #11 (Jack Point, non-silvery).

The silvery-leafed plants and the non-silvery-leafed plants allowed to self-fertilize yielded all silvery and all green progeny respectively, indicating they are homozygotes. These data agree with the results discussed above. However, the results of the cross- fertilizations are not as expected. The cross, #25 non-silvery (9) x

#25 silvery (c?) yielded only non-silvery-leafed offspring instead of the heterozygous silvery-leafed offspring that were expected. These non-silvery-leafed plants are probably the results of accidental selfs. Since emasculating and cross-pollinating the flowers of population #25 is so difficult, accidental selfing can probably occur quite easily. If this is the case, the other crosses involving seed parents from population #25 are also suspect and the silvery progeny from these crosses may also be the result of accidental selfing. These suspicions were confirmed in the next generation when all of the F., progeny resulting from crosses where a silvery- leafed plant was used as the original seed parents were silvery.

This indicates that all the silvery F^ progeny were the result of accidental selfing. The F^, and S^ progeny are summarized in

Table VII, Table VII

SP and Fp progeny of the crosses involving the silvery-leaf character

of S2 & F2 # of Sg. & F2 individuals Origin of and F.^ plants families Silvery Green

Population #25; silvery, selfed 12 1,203 0

Population #25; non-silvery, selfed Ik 0 1,156

#25 silvery (9) x #25 non-silvery (c?) 5 765 0

#25 non-silvery (9) x #25 silvery (d1) 8 0 630

#25 silvery (9) x #11 non-silvery (c?) if 220 0 - 30 -

The one cross that appears to have been successful is

#11 non-silvery (9) x #25 silvery (cf). The flowers from population

#11 (Jack Point) can be emasculated and cross-fertilized quite successfully as shown in the spotted-leaf experiment. The 7 green- leafed progeny in the F^ generation (Table VI) may be the result of a cross with a heterozygous silvery plant but since all of the self- fertilizations (accidental or otherwise) seemed to indicate only homozygotes, it is more probable that these progeny resulted from the accidental selfing of one or more flowers of the seed plants.

However, the 15 silvery offspring of this cross can only be heterozygotes and the result of successful crossing since population

#11 contains no silvery-leafed genes. These silvery offspring were initially scored as 'intermediate1 since they were not as clearly silver as the plants resulting from the self-fertilizations (Fig. 9).

This made it attractive to postulate that heterozygotes could be distinguished from homozygotes by the degree to which the silvery

character is expressed. However, when the F2 progeny of these

•intermediate' F^ plants were grown, the silvery plants did not segregate into the two categories, silvery and intermediate as expected, but stayed mostly in the intermediate category with almost no plants showing the clearly silvery leaf of the population #25 plants. This would indicate that the 'intermediate' expression is due to the modification of the silvery gene by the population #11 genotype rather than to the effects of heterozygosity.

The F2 progeny of the #11 green (9) x #25 silvery (cf) cross are summarized in Table VIII. - 31 -

Fig, 9 Silvery, green and •intermediate1 plants Table VIII

Fp progeny of the #11 green (?) x #25 silvery (cf-) cross

# of F2 individuals Y2 . - x Phenotype of F, # of F. families A vx a,I,; Silvery Green

Non-silvery 7 h 932

Silvery (intermediate) 13 1,026 347 0.05$ P=0.9-0.5 deviation not significant 33 -

The non-silvery plants bred true as expected. The If silvery plants were probably contaminants from neighbouring silvery families.

The silvery plants segregating into an approximately 3*1 ratio indicates that the silvery-leafed character is a simple dominant.

Discussion

The silvery-leaf polymorphism found on Carlos Island is con• trolled by a single gene with a dominant allele for presence of the silvery-leafed character and a recessive allele for the absence of it. The dominant allele is very strongly expressed and very obvious in plants grown under artificial conditions. Unfortunately however, the presence of the silvery-leafed character is difficult to identify in nature.

So far, this silvery-leaf polymorphism has not been found in any populations other than on Carlos Island, Controlled crossing with plants grown from seed from Jack Point indicate that other genotypes can modify the expression of this gene. - 34 -

Chapter k Flower Colour Mutants

Introduction

The flowers of both the large and small-flowered £_ parviflora are blue with varying amounts of magenta and white in the upper lip.

The varying intensity of pigment from plant to plant and in different parts of the flower produces different shades of colour ranging from light blue to dark purple. In his original description, Lindley

(1827) describes as "one of the most beautiful hardy annuals with which we are acquainted, covering the ground with a carpet, as it were, of blue, purple and white, during the months of June and July".

All of the populations studied had normal blue flowers, however, two populations from Vancouver Island also contained plants with atypical flower colour. Some of the plants from Elk Falls, a population of large-flowered Cj, parviflora, had pink or magenta flowers (Fig. 10) and some of the plants from Mt. Douglas Park, a population of small-flowered £. parviflora, had flowers that were completely white (Fig. 11).

These two unusual flower colour mutants had not been reported in the area of this study but St. John (1956) reports these colours to be present in two small-flowered ("parviflora") populations in southeastern Washington and formally recognizes them as formae.

Forma alba English is described as having white corollas and forma rosea Warren is described as having rose-mallow corollas.

Harborne (1967) states that "a general ... characteristic of white flower mutants of coloured forms is their unthriftiness as plants". This was clearly true of the white flowered Collinsia. - 35 -

Fig. 11 An albino flower from the Mt Douglas Park population - 36 -

The white-flowered population, although it survived well under ideal culture conditions was more drastically affected by aphid attack, fungus infection and drying out than the normal blue-flowered populations* It is interesting to note that aphids damaged the white flowers quite badly, often hampering successful cross-fertil• izations but did not seem to attack those flowers containing antho- cyanins, indicating perhaps that one of the functions of anthocyanin pigments in parviflora is protection from insect pests or that the character is linked with another chemical character.

It was observed that plants with blue or magenta flowers often had purple pigment on the underside of leaves. This pigment also tinged the upper surfaces of leaves in all except the white-flowered plants as the plants aged (Fig, 12), Pigment production could be considerably speeded up by increasing the light intensity or by an aphid attack.

Greater leaf pigment production under higher light intensities is probably related to the fact that light has been shown to be important for initiating or stimulating flavonoid synthesis

(Harborne, 1967),

Injury can also stimulate anthocyanin synthesis or even initiate pigment formation in tissues that usually do not contain any antho• cyanin colour (Bopp, 1959), This was the case when aphids infested a population. Then purple pigment was produced in the leaves even in the seedling stage.

Plants with white flowers did not produce any anthocyanin and when the plants aged or were infected by aphids or fungus the leaves

turned yellow (Fig, 13), - 57 -

Fig, 13 The Mt, Douglas Park population with yellow leaves — 38 —

Through crossing experiments, I attempted to determine the inheritance of these flower colour mutants in £. parviflora.

Materials and Methods

Population #2 was used as the source of white-flowered plants.

It was grown from seed collected from white-flowered plants at Mt.

Douglas Park (Fig. 1). Population #17 grown from seed collected in

Elk Falls (Fig. l) was the source of both blue and magenta-flowered plants. In addition two more populations of blue-flowered plants were used; population #9 (Nanoose Hill) and population #11 (Jack

Point). Some plants from each population were allowed to self for two generations and others were used in the crosses..

The following crosses were set up between blue and white- flowered plants:

#9 blue (9) x #2 white (cf)

#2 white (9) x #9 blue (cf)

#11 blue (?) x #2 white (cf)

#2 white (9) x #11 blue (cf)

The crosses between blue and magenta-flowered plants were:

#17 blue (9) x #17 magenta (cf)

#17 magenta (9) x #17 blue (cf) 39 -

And the crosses between white and magenta-flowered plants were:

#2 white (?) x #17 magenta (cf)

#17 magenta (?) x #2 white (c?)

In addition, two sets of crosses from the flower size study

showed flower colour segregations in the F2 generation and so were used to supplement the data from the above-mentioned crosses* In the crosses #17 (?) x #22 (cf); #22 (9) x #17 (cf), both blue and magenta-flowered plants from population #17 were used so that in

the F2 generations some of the families showed a segregation for flower colour*

The other set of supplementary data came from the cross

#22 (9) x #9 (c?)* This was also a cross used in the flower size

study* Both populations were blue-flowered* Some of the F2 families from this cross showed segregation for blue and white flower colour*

This was completely unexpected since no other cross or self-fertil• ization of either of these two populations produced any white-flow• ered plants* One possible explanation for this segregation is that there was a mutation in one of the parent plants which was passed on through this particular cross. Another is that the white flower gene is present in one of the populations in such low frequencies that it is rarely in its homozygous condition and therefore rarely detected. If this were the case, one of the original parents of the cross would have been heterozygous for the white flower gene*

Whatever the reason, the white flower gene was present in seven of

the F2 families and so could be used in determining its inheritance pattern* ko -

The cross-fertilization methods, growing conditions and scoring methods were basically the same as for the studies discussed in previous chapters. In most crosses more than one plant was used as pollen parents and seed parents. However, in crosses involving the Elk Falls population, only one pollen parent and one seed parent were used in each case, since the Elk Falls flowers are very large and easy to manipulate* Seeds from the crosses and selfs were collected, grown, scored for flower colour and tagged* Seed from each F^ and S^ plant was then collected and grown separately* The

F2 and S2 families were scored separately to determine the F^ and S^ genotypes and eliminate any contaminants and then the frequencies were totalled to get a more accurate ratio*

The F2 generation of the blue and white coloured crosses was attacked by aphids and most of the plants were killed in the seedling stage* However, a tentative count was made without flowers, based on the observation that white-flowered plants turn yellowish and blue- flowered plants turn purplish when damaged by aphids* The plants that remained green were not included in the count*

Results

The results from the plants that were allowed to self are summarized in Table IX*

The plants from all the populations appear to breed true for flower colour* This large amount of homozygosity is expected in populations #9 and #11 since the magenta and white flower colour mutants are probably not present. The high frequency of homozygosity Table IX

S1 and S2 progeny of the populations used in the crosses involving flower colour

Population Flower colour # and flower colour # and flower colour

(allowed to self) of parent plants of individuals of S2 individuals

#2 Mt. Douglas Park White 20 plants - all white ifl7 plants - all white

#9 Nanoose Hill Blue 13 plants - all blue i+Ok plants - all blue

#11 Jack Point Blue 19 plants - all blue 1,169 plants - all blue

#17 Elk Falls Magenta 3 plants - all magenta (S^ died before seed set)

#17 Elk Falls Blue 30 plants - all blue 56 plants - all blue - 2f2 -

in population #1? is not surprising either since the original sample was very small, only four or five plants. And the white-flowered plants from population #2 appear to be homozygous recessive. The fact that all of the plants left to self were homozygous indicates that all or most of the plants used in the crosses were probably also homozygous. Crossing data supports this in all of the crosses except

#22 blue (9) x #9 blue (cf) which was discussed above.

The offspring of the cross between blue and white-flowered plants were all blue except for 3 white-flowered plants which resulted from the cross #2 white (9) x #11 blue (cf) and these were probably the product of an accidental self of the #2 white seed parent rather than

a cross with a heterozygous population #11 plant. In the F2 gener• ation there was segregation for both blue and white-flowered plants.

The results of these crosses are summarized in Table X.

The total F2 score for all of these crosses is 92 blue : 32 white, an almost perfect 3:1 ratio. This simple Mendelian ratio makes it attractive to postulate a single gene but the data were

obtained from a tentative count of dying seedlings and hence the

experiment should be repeated to substantiate this ratio.

The other cross resulting in the segregation of blueand white-

flowered plants was #22 blue (9) x #9 blue (cf). In this cross,

seven F2 families showed segregation into blue and white-flowered

progeny but they did not conform to the expected 3 blue : 1 white

ratio. The total score for these families is 403 blue : 41 white

(Table XI). Although these data do not fit a common Mendelian ratio

they are more reliable than the 3:1 ratio obtained from the dying Table X

and F2 progeny resulting from crosses between blue-flowered and white-flowered plants

Cross # of F, individuals # of F~ individuals Blue White Blue White

#9 blue (9) x #2 white (cf) 9 0 15 k

#2 white (?) x #9 blue (c?) 7 0 9 2

#11 blue (?) x #2 white (c?) 31 0 36 10

#2 white (9) x #11 blue (<*) 11 3* 32 _l£

Total 58 3 92 32

•accidental selfs Table XI

F2 progeny of the #22 blue (9) x #9 blue (cr) that showed segregation for flower colour

Colour of the F, plants # of individuals in each family which produced each family Blue White

1. Blue 89 5

2. Blue 32 9

3. Blue 59 6

4. Blue 61 8

5. Blue 70 5

6. Blue 38 2

12. Blue 6

Total 403 41 - 45 -

seedling count discussed above* It is reasonable to assume that this seedling count underestimated the number of blue-flowered progeny since only the unhealthy seedlings whose leaves had turned colour were scored* And since white-flowered plants are less hardy and more severely affected by aphids, most or all of the healthy and still green seedlings omitted from the analysis were probably blue-flowered plants* Had they been scored, the final ratio would probably have shown an unexpectedly high proportion of the dominant blue-flowered

F2 offspring.

In the crosses between blue and magenta-flowered plants the F^ plants were all blue-flowered, indicating that blue flowers are

dominant over magenta flowers as well as white. In the F2 generation the families showed segregation for blue and magenta flowers. How•

ever, the ratio was again an anomalous one, 50 blue : 6 magenta.

These results are summarized in Table XII. Thus the magenta flower

character, like the white flower character, is transmitted at a

frequency that is much lower than expected for simple single gene

inheritance.

The cross between magenta and white-flowered plants produced

all blue-flowered F.^ progeny except for one white contaminant proving

that white and magenta flower colour are not allelic. And the F2

generation showed segregation for blue, magenta and white flowers

(Fig. 14). The results are summarized in Table XIII. Table XII

F^ and F^ progeny of the crosses between blue-flowered plants and magenta-flowered plants

# of F2 individuals # and colour of (by individual family) Pollination regime -L F individuals Blue Magenta

1 0 #17 blue (9) x #17 magenta (cf) 11 blue 5 0 1 0

#17 magenta (9) x #17 blue (cf) 3 blue

#22 blue (9) x #17 magenta (

6 3 #17 magenta (?) x #22 blue (cf) 2 blue 20 1

Total 17 blue 50 - 47 -

Fig, 14 The F2 generation in the cross between white and magenta-flowered plants, showing segregation for the three flower colours Table XIII

F^ and F2 progeny of the crosses between white-flowered and magenta-flowered plants

# and flower colour of # of F2 individuals Pollination regime the p in^yi,^^ Blue Magenta White

#17 magenta (?) x #2 white (cf») 10 blue 18

#2 white (?) x #17 magenta (c?) 65 blue 148 16 12 (1 white)*

Total 75 blue 166 18 13 (1 white)*

•accidental self - 49 -

Discussion

These data suggest that at least two loci are involved, with one gene (A) regulating the production of magenta pigment from a colourless precursor and a second gene (B) regulating the production of a blue pigment from the magenta one* i.e.

colourless precursor ^ene A—> magenta gene B—^ blue

If this is the case, the white-flowered plant has the genotype aaBB and the magenta-flowered plant has the genotype AAbb* The hybrid is AaBb and blue* And the blue-flowered plants used in the crosses were all AABB*

However, using the above hypothesis, the F2 ratio for the crosses between blue-flowered and white-flowered plants would be expected to be 3 blue : 1 white. But the actual ratio was 403 blue : 41 white.

The crosses between blue and magenta would also be expected to yield an F-> ratio of 3:1 instead of the unusual 50 blue : 6 magenta ratio that was actually produced* This very low frequency of the recessive allele also occurred in the crosses between white and magenta-

flowered plants. The expected ratio would be 9 blue : 3 magenta :

4 white. The actual ratio was 166 blue : 18 magenta : 13 white.

One of the simplest ways to explain these data is to postulate

that only two genes are actually involved but that there is a

selective pressure against the recessive alleles in the formation of

gametes or against the homozygous recessive condition in the zygote

or young seedling. - 5© -

This is not the only possible explanation for this unusual ratio. It may be a much more complex system involving more than two loci and various gene interactions. But a system of three loci can• not explain all of the information and there are not sufficient data to postulate more genes. Therefore at this point the simplest hypothesis is the one discussed above. However, it must be further tested to determine whether or not selection against the recessive alleles is actually taking place. If the hypothesis proved false, more complex genetic determinants would have to be investigated.

Although the inheritance patterns for the flower colour poly• morphisms have not been conclusively determined, this study has provided a feasible working hypothesis as a basis for initiation of further studies into the problem. - 51 -

Chapter 5 Inheritance of Flower Size

Introduction

Flower size is the main character used to separate £. parviflora and C. grandiflora in most floras and in Newsom's (1929) monograph of the genus. The original descriptions (Lindley, 182?) do not give any actual measurements but later authors divide the two species into two very precise and measurable size classes. However, they do not always agree on the range in flower size of each species, the degree of overlap between the two or the point at which the two species are separated. Table XIV summarizes how various authors have treated this character.

The dividing line between grandiflora and parviflora has been drawn seemingly arbitrarily at different flower sizes by different authors, Abrams (1951) includes in C_, parviflora those plants with corollas less than 10 mm long whereas Davis (1952) states that C. parviflora has corollas less than 6 mm long and £• grandi• flora has corollas between 6,5 and 9 nim long.

Only Peck (1961) and St. John (1956) have formally recognized any region of overlap between the two species in their keys and descriptions. However, Newsom (1929) does state in her text that

"in studying these two closely allied species, one finds a continuous series of intergradation from the largest-flowered grandiflora to the smallest-flowered parviflora". Other authors, notably Gilkey and Dennis (1967), make the two size classes appear to be quite distinct.

This is not the case in nature. In fact there seems to be a more or less continuous range in flower size among the populations Table XIV

Corolla length classes assigned to C. parviflora and C. grandiflora by various authors

C. grandiflora C. parviflora

„ . » Total corolla lengte h (mm) Author Region of flora 1 2 3 k 5 6 7 8 9 10.11 12 13 14 15 16 17 18 19

1 1 i 1 Peck (1961) Oregon I—. 1 Davis (1952) Idaho h •• ' 1 St. John (1956) S.E. Washington 1 1 Henry (1915) Southern B.C. +• 1 ' ' Anderson (1961) Alaska *" ^ Munz & Keck (1959) California "H 1 ' Lindley (1827) Orig. descript. no size estimate Newsom (1929) Monograph \ T ' ' Hitchcock et al. (1959) Pacific N.W. I- 1 ' f Piper (1906) Washington ' ^ ' ' Piper & Beattie (1915) 1 1 I 1 Gilkey & Dennis (1967) Pacific N.W. I-i Abrams (195D Pacific States J 1 1 - 53 -

observed in this study. And many of them occupy the regions of division chosen by various authors and so are difficult to assign confidently to one species or another.

Because of the importance placed on flower size as a key character and because of the differing opinions among various authors using this character, it was of special interest to examine variation in flower size both among and within populations in this study and to determine the genetic mechanism controlling it.

Materials and Methods

Flower size varies considerably among populations of C. parvi• flora in southwestern British Columbia and northwestern Washington.

For example, flowers from the Lindeman Lake population are usually

4-5 mm whereas flowers from Elk Falls are about 10-1? mm. However, the flower size within any particular population is usually relatively uniform and populations grown from seeds collected in different areas can often be visually distinguished merely on the basis of flower size. For this particular study five populations were chosen on the basis of flower size, ranging from the population with the largest observed flowers to the population with the smallest observed flowers and with each population visibly different from the others in the size

of its flowers (Fig. 15). Three of these populations fell within the

C. grandiflora range according to Hitchcock and Cronquist (1973) and

two fell within the C. parviflora range. See Table XV.

The flowers of the Lindeman Lake population were very small and

difficult to work with. Emasculation of the tiny buds was not - 54 -

Fig. 15 The variation in flower size between the five populations used in study. From left to right: Elk Falls, Jack Point, Nanoose Hill, Botanie Valley, Lindeman Lake Table XV

Populations of C. parviflora used in the study of flower size variation

Species according to Total corolla Population Locality Hitchcock & length number Cronquist (1973)

10 17 mm Elk Falls 17 C« grandiflora

8 10 mm Jack Point 11 C, grandiflora

7 8 mm Nanoose Hill 9 C. grandiflora

6 7 mm Botanie Valley 22 C. parviflora

k 5 mm Lindeman Lake 6 C, parviflora - 56 -

possible and so they were only used as a pollen parent. Plants from population #6 were crossed with plants of each of the other populations.

The Botanie Valley population had slightly larger flowers than those from Lindeman Lake and so was less difficult to work with.

Reciprocal crosses wereset up between plants from this population and plants from each of the "(_. grandiflora" populations. Some plants from each population were also allowed to self. Seeds from the cross-fertilizations and self-fertilizations were grown to pro• duce the F-^ and S^ generation which was in turn allowed to self to

produce the F2 and S2 generation. A sample of flowers was measured for each of the .original parental populations as well as each of the

Fl' Sl' F2 ajx^i S2 Populations except for the S^ generation of popul• ation #11 (Jack Point).

To minimize the effect of differing flower angles the measure• ment taken was from the tip of the keel to the top of the saccate or gibbous swelling at the base of the corolla tube (See Fig. 16).

Fig. 16 A Collinsia flower showing the angle at which measure• ments were taken

The measurements were taken to the nearest 0.5 mm. These were later rounded off to the nearest mm in order to simplify the graphing.

Samples of three flowers per plant were taken (whenever possible) from up to twenty plants in each parental, S^._ and F^ population. In 57 -

the and generation three flowers (whenever possible) per plant were taken from up to ten plants per family and up to nine families from any particular self or cross.

These samples were analyzed and compared by totaling the number of flowers in each size class (to the nearest mm) in the sample and converting them to percentages. The percentages were then graphed.

In addition to each graph, a table was set up indicating the mean flower size and the sample size of each population involved.

One of the crosses set up for flower colour (population #2 x population #17) also showed interesting results as far as flower size was concerned. Population #2 (Mt, Douglas Park) had "parviflora" flowers and population #17 (Elk Falls) had very large "grandiflora" flowers. The hybrids were obviously intermediate. Samples were measured, analyzed and compared in the same way as the crosses dis• cussed above. But because this was not originally designed as a cross to determine the inheritance of flower size there is no sample for the original parental population from Mt, Douglas Park,

Results

The graphs for flower size of the five populations showed a certain amount of overlap between the populations and some shifting of the histograms from one generation to the next but the peak of each histogram was clearly separate from all the others (Figs, 17,

18, 19).

The mean flower sizes were also different for each of the populations (Tables XVI, XVII, XVIII), The shifting of the histograms Figur< 17 i Graph c omp arinj jri c wer sizes < r the fiv e populations in 1th e i "irst (parenta] /) a•eneratio n

•

o 8u

It £_ / o / •3 c / \ / / \ J re1 \ #22 i 4-= f < +• » «w 40 1

M #13 »

2 4 8 1() 12 14 1 6

flowe.c tii :e • ( rar

J. 11II L 1 Table XVI

Mean flower sizes and sample sizes of the populations graphed in Fig, 1?

e Z Population x flower size (# Qf pla^f g *f flowers)

Lindeman Lake 4»0 mm 5 15

Botanie Valley 4*5 mm 9 2?

Nanoose Hill 6,0 mm 5 15

Jack Point 7,0 min 6 18

Elk Falls 10,5 mm 18 53

Table XVII

Mean flower sizes and sample sizes of the populations graphed in Fig. 18

_ , . . - . Sample size Population x flower size (# Qf plants) (# of flowere)

Lindeman Lake 3.0 mm 15 45

Botanie Valley 4»0 mm 9 26

Nanoose Hill 5*0 mm 11 33

Elk Falls 13.0 mm 17 40 Figure 19 Graph comparing flower sizes of the five populations in the

flower size (mm) Table XVIII

Mean flower sizes and sample sizes of the populations graphed in Fig. 19

- „n . Sample size Population x flower size (# Qf plante) (# of flowers)

Lindeman Lake 2.5 mm 50 121

Botanie Valley if.5 mm 72 188

Nanoose Hill 6.0 mm 72 186

Jack Point 7.5 mm 40 102

Elk Falls 11.5 mm 60 164 from one generation to the next may simply be due to differences in the age of the plants when the flowers were measured. It was ob• served, particularly in population #17, that the plants produced smaller flowers as they aged. These shifts could also be due to environmental factors. However, since the plants were all grown under uniform growth chamber conditions, the environmental effects would be minimal.

The crosses involving the Lindeman Lake population (#6) as a pollen parent were not very successful. Very few seeds were set and most of those seemed to be contaminants since the supposed 'hybrids' resembled their seed parents. This was largely a technical problem since the flowers from this population were very small and had very little pollen. Most of the pollen was usually lost in the attempt to transfer it onto another stigma. However, at least one or two pro• geny appear to have been genuine hybrids because in the F^ generation one family from the cross #11 (9) x #6 (6*) had flowers of an inter• mediate size (See Fig, 20) and one family (consisting of a single plant) from the cross #9 (9) x #6 (cf), had flowers in the size range of population #6, i.e. 2,5 - 3-5 mm.

The crosses using the Botanie Valley population (#22) for the

"parviflora" parent were more successful. Hybrids were readily

formed and the F.^ and F2 hybrid peaks were intermediate to the parental peaks in most cases (Figs, 21, 22, 23), The mean flower sizes of the hybrids were also intermediate between the parents

(Tables XX, XXI, XXII), And the Fg generation showed a greater

variation in flower size than the Fn generation, suggesting segreg- Figure >

(5ra i )hL C 0m pa rini tti e tlowe r sizei 3 <> f Lindeman h£ ike (# 6 ) > i J ack P 01 nt L]• I»opu la t ions i •Vi cm : )( ) with a ] L

-

r> 0 0

n (1) 1 \ ft hiill o ow i cr V nv 1 / i i j 5 1 c 0

/

20 •\ %

••'

_ .• 4 6 10 12 14 16

i'lowe r siz 3 {[mn L ) Table XIX Mean flower sizes and sample sizes of the populations graphed in Fig, 20

Sample size Population x flower size (# of plants) (# of flowers)

Lindeman Lake parental if,0 mm 15

Jack Point parental 7,0 mm 18

Possible F2 hybrid 6,0 mm 10 21 Figure 21 Graph comparing the flower sizes of the hybrids between populations from 100 Botanie Valley (#22) and Nanoose Hill (#9) with their parental populations

flower size (mm) Table XX

Mean flower sizes and sample sizes of the populations graphed in Fig. 21

Sample size Population x flower size (# of plants) (# of flowers)

Botanie Valley parental if*5 mm 9 27

4»5 mm 53 Fx hybrid 18

5.5 mm 160 380 F2 hybrid

Nanoose Hill parental 6.0 mm 5 15 Figure 22 Graph comparing the flower sizes of the hybrids between populations from 100 Botanie Valley (#22) and Jack Point (#11) with their parent populations

80

CO i - o 60

I H (0 ON +> VC O +» 1

© k0 #22 parental

20

8 .0 ] ll> Ik

im) -

... Table XXI

Mean flower sizes and sample sizes of the populations graphed in Fig, 22

Sample size Population x flower size (# of plants) (# of flowei

Botanie Valley parental 4,5 mm 9 27

Fx hybrid 5,5 mm 18 49

F2 hybrid 6,5 mm 120 290

Jack Point parental 7,0 mm 6 18 Figure 23 Graph comparing the flower sizes of hybrids between populations from

flower size (mm) ^TJTTJTTlTtl IMIIIII H-H-l 1111111M1III III M HH+H^B Table XXII

Mean flower sizes and sample sizes of the populations graphed in Fig. 23

- „ . Sample size Population x flower size (# Qf plants) (# of fi0Wers)

Botanie Valley parental 4»5 mm 9 27

F1 hybrid 8,5 mm 11 33

F2 hybrid 7.5 mm 107 187

Elk Falls parental 10.5 mm 18 53 - 73 - ation for the genes determining flower size (Figs, 21, 22, 23), This would be expected if flower size is polygenically controlled. The cross between population #17 and population #22 produced similar results (Fig, 24; Table XXIII),

These results indicate that the character of flower size is controlled by polygenic or quantitative inheritance though it is not clear how many genes are actually involved.

Discussion

The graphs (Figs, 17, 18, 19) confirmed previous observations that flower size varies between populations and yet is relatively uniform within any particular population. The amount of overlap between the different populations, especially between "grandiflora" and "parviflora" populations weakens the case for using flower size as a key character especially considering that this was a relatively small sample of populations chosen particularly on the basis of their clearly different flower size distributions, A more extensive col• lection of populations would undoubtedly blur the taxonomic dis• tinctions even more. In addition, the "grandiflora" and "parviflora" populations can interbreed quite readily. This clearly indicates that they are not good biological species in the sense of Mayr (1957) and that there is only one biological species, £, parviflora.involved. However, if the morphological characters of £, grandiflora and C, parviflora are sufficiently distinct and different, they could be separated at the level of taxonomic species as distinguished from biological species F ignr 9 >J«•

1 Graph comparing the flowei size s of ]K fbr ids 3 1betwee n 1copulatio n B fron l n.0 0 J Mt • Dougla (#2 ) sine1 Elk F a llo FX/; witl r ]jarem ; popuiax-xons

80

r /fr-tt s r-l «H >n 1 H / \ l OS +> • O *» i <*

F hybrit 1 o 40 l / / \ #17 ]parenta l \ F \ i \ 2 / \ / y 20 I \ / / \ T VT / \ / \ H N "••-...\ V- c> 6 \3 it 12 14 Lc

1 lc •wer siz [mm)

1 _ «L _ Table XXIII

Mean flower sizes and sample sizes of the populations graphed in Fig. 24

Sample size Population x flower size (# of plants) (# of flowers)

47 Mt. Douglas Park S1 4.5 mm 18

F^ hybrid 9.0 mm 20 70

388 F2 hybrid 7«5 mm 154

Elk Falls parental 10.5 mm 18 53 - 76 -

by Cain (1954) and Grant (1963» 1971)• This cannot be done with the character of flower size which shows a continuous variation, however other morphological characters will be examined in Chapter 7 in order to determine whether the recognition of two or more taxonomic specieB can be justified. - 77 -

Chapter 6 Chromosome Numbers

Introduction

Garber and his students have done extensive work on the chromo• somes of Collinsia and their studies indicate that with one exception, the species all have seven bivalents at metaphase I (Garber, 1956,

1958a, 1958b; Ahloowalia and Garber, 1961; Garber and Dhillon, 1962,

Hayhome and Garber, 1968; Garber, 1974)• The one exceptional species is C, torreyi with a count of 21 bivalents, making it a hexaploid. It was at first considered the "first polyploid to be encountered in the genus" (Garber, 1958b) but later was said to have been "erroneously identified as a member of Collinsia (since) no tetraploid species has been found in the genus" (Garber, 1974)• This casts some doubt on the identity of the plants for which Garber has reported chromosome numbers, especially since there are no voucher specimens* However,

Taylor and Mulligan (1968) in their Flora of the Queen Charlotte

Islands, give the chromosome number of £, parviflora as n=14, making

it a tetraploid,

Garber's chromosome counts for C, parviflora (1956) and C^ grand•

iflora (1958b) are both n=7 but the parviflora count came from only

two pollen mother cells and the grandiflora count appears to have

come from only one plant, Taylor and Mulligan took their count from

a population on Haida Pt,, Graham Island but Garber made no mention

as to the source of his populations.

Because of these conflicting reports I examined six populations

with different flower sizes to determine their chromosome number.

The objectives were to determine if there were differences in number

in different populations, if both tetraploids and diploids existed in - 78 -

this area of study, and whether small and large-flowered populations differed in chromosome number.

Materials and Methods

Plants were grown from seed collected at the following localities (See Fig. 1): it Population #6 - Lindeman Lake

Population #22 - Botanie Valley Population #9 - Nanoose Hill Population #25 - Carlos Island Population #11 - Jack Point

Population #17 - Elk Falls

Buds were fixed in a 6:3?2 solution of ethanol, chloroform and propionic acid and then stained in alcoholic hydrochloric acid-carmine

(Snow, 1963). These buds were then dissected, the anthers removed and

squashed. Since the anthers mature at different times and all of the

cells in a single anther do not divide synchronously, a bud with

tetrads often also contained a few pollen mother cells at late pro•

phase or metaphase I. The chromosomes tended to be quite sticky and

were difficult to count but at least one cell with distinguishable

chromosomes was found and photographed for each population. Vouchers

are deposited at the University of British Columbia.

Results

The following counts were obtained from the populations studied:

Population #6 - Lindeman Lake 2n = 14 II

Population #22 - Botanie Valley 2n = 14 II - 79 -

Population #9 - Nanoose Hill 2n = 14 II (See Fig. 25)

Population #25 - Carlos Island 2n = 14 II (See Fig. 26)

Population #11 - Jack Point 2n = 14 II

Population #17 - Elk Falls 2n = 14 II

Discussion

All of the populations surveyed for chromosome number were tetraploids with a count of 2n = 14 II. This agrees with Taylor and Mulligan's (1968) count for C_. parviflora from the Graham Island population and suggests that C. parviflora is tetraploid in British

Columbia.

None of the populations studied had a chromosome number of n=7

as reported by Garber (1956, 1958b). It is possible that C^ parvi•

flora consists of both diploid and tetraploid populations and Garber

obtained his plants from diploid populations. However, it is also

possible that C^ parviflora is a strictly tetraploid species and the

material from which Garber obtained his counts was misidentified as

parviflora and £. grandiflora. It is unfortunate that he does

not state the source of his seeds.

Both large-flowered and small-flowered populations had a chromo•

some number of 2n = 14 II, supporting the inclusion of both large and

small-flowered plants in the same species. Since large and small-

flowered plants formed fertile hybrids, this result was not unexpected. - 80

Fig. 25 A pollen mother cell from the Nanoose Hill population showing a chromosome number of 2n = 14 II (mag, x 1,650)

Fig, 26 A pollen mother cell from the Carlos Island population showing a chromosome number of 2n = 14 II (mag. x 1,900) - 81

Chapter 7 Morphology and Taxonomy

Introduction

It was established in Chapter 5 that £. grandiflora and

C. parviflora are interfertile and therefore not separate biological species in the sense of Mayr (1957) and Grant (1963)» However, this is not sufficient reason to ignore the nomenclatural distinction between the two. It may be useful in practical taxonomic work to consider them to be two taxonomic species (Grant, 1971) which express morphological differences rather than crossability or interfertility relationships. The following morphological study was conducted in an attempt to determine whether the "grandiflora" and "parviflora" groups are sufficiently different and distinct to be considered separate taxonomic species.

Materials and Methods

Sixteen populations were grown from seed collected from the

following localities (See Fig. l):

Vancouver Island. B. C.

Thetis Lake - population I - population II

Mill Hill - population I - population II

Mt. Douglas Park - population I - population II

Nanoose Hill - population IV - population III - 82 -

Nile Creek

Little River

Crofton - population I - population II

Rathtrevor Park

Sechelt Peninsula. B. C.

Lund

Irvine's North

Washington. U.S.A.

Anacortes

In addition five of the populations used in the genetic studies were used in the Sg generation:

#6 Lindeman Lake, B. C.

Vancouver Island, B. C.

#2 Mt. Douglas Park - (population III)

#9 Nanoose Hill - (population I)

#11 Jack Point

#17 Elk Falls

All of the above populations were planted in shallow flats in growth chambers and allowed to reach maturity, at which time veget• ative and floral characters were measured.

The vegetative and floral characteristics of £• grandiflora and

C. parviflora. especially those that reportedly differed between the two groups, were summarized on a data sheet (Table XXIV). Each 83 -

Table XXIV

Data Sheet for Population

Stem erect, ascending, other branched, unbranched glandular, puberulent, glabrate, other tall

Leaves oblong, ovate, spatulate, other obtuse, acute, other serrulate, entire, entire-revolute, other glabrate, puberulent, glandular, other sessile, sub-sessile, petiolate upper whorls becoming linear

smaller bracteolate in i purple underside ______long; wide

Flowers flowers in whorls rarely solitary, commonly solitary below, never solitary

Pedicels puberulent, glandular, glabrate, other times as short as flowers long

Calyx glabrate, puberulent, glandular, other membranous below ______scabrous-margined ______times length of corolla long lobes: subulate, linear-lanceolate, other acuminate, other wide - 84 -

Corolla strongly declined, declined, almost erect forms 0 angle with pedieel long upper lips colour ______long sinus deep lobes: obovate, spatulate, retuse, dilate, crisp-crenulate, recurved, erect, other wide

lower lip: colour wide

corolla tube: colour long saccate, gibbous, other sparsely bearded, glabrous, other

Comments: - 85 - population was then surveyed for the presence or absence of each possible character. Corolla size was measured as the total length of the corolla rather than the measurement used in Chapter 5 in order to make the measurements comparable to those of other workers. The data were then summarized in Table XXV to include all those characters that showed variation among the different popul• ations. In this form the data could be examined to determine any patterns or obvious differences which could be used to divide the populations into two or more groups.

Results a) Stem In the populations studied, the stem height ranged from 2.4 cm (Rathtrevor) to 25 cm (Nanoose Hill I). Newsom (1929) describes C. parviflora stems as ascending or erect as compared to £j, grandi• flora stems which are almost always erect. However, the populations in this study cannot be separated on this basis since all plants were erect. They were also all puberulent. The only stem character that varied was branching. Most populations showed branching but the absence of branching could not be correlated with any other character.

b) Leaves The greatest amount of variation within and between populations was found in the leaves. Leaves varied in size, shape, leaf margin, pubescence, pigmentation and length of petioles (See Figs. 27, 28). However, this variation was often as great or greater within a -86-

Table XXV Summary of the variation among the populations studied

. . • • A. «_ __> A* • t. I_l i_l 1—4 • I. c^-t t<+ mH-ft* P* P4 H «•*** «-t* Ua PT* FT I—«• rr<|0&tJctHOOI»rtHcHJB • • • H CD CD P F- o a.|3*HHi,M»o*+'co cu oo H ft cr H Hrjfcj O <+_ ct- d- O H POO O O O _ H- H- ffl O O O W CD OJ 5 P CD *t> a iHDoo»i»OrkiiiB £ 0 P H* £ M - O CD CD H' P P ct- 4 CD CD 1/2 a corolldeclinea dlengt ( h 45°) + * + + + + corolla almost erect tube saccate + + + + + + + + + + + + + + + tube gibbous + + + + + + + + + corolla limb ^. tube + + + + + + + + + + + + + ++ + + + + + + + corolla limb ^ tube + + + + + corolla k mm + + + + + + + + corolla 5 mm + + + + + + + + + + + •»•+ + corolla 6 mm + + + + + + + + +•+ + + + + + + + corolla 7 mm + + + + + + + + + + + + corolla 8 mm + + + + corolla 9 mm corolla 10 mm + corolla 7" 10 mm - 15 mm - 87 -

Fig, 27 The leaf characters of the Botanie Valley population

Fig. 28 The leaf characters of the Jack Point population - 88 particular population than "between populations and it is not possible to use leaf characters as a means for separating the populations into different taxa. c) Flowers

The number of flowers per whorl has been used as a diagnostic character separating the two species. Newsom (1929) describes

C. parviflora as having "2-5 flowers in a whorl above, usually solitary below". By contrast, she describes C. grandiflora as having

"flowers 3-7 at a node, rarely solitary". As can be seen in Table XXV, all populations except Nanoose Hill IV had flowers "usually solitary below". As for the upper whorls, all populations had flowers in 2's and only three populations had whorls with more than 5 flowers.

These were Mt. Douglas Park III, Nanoose Hill I and Elk Falls and include both "parviflora" and "grandiflora"-size flowers.

d) Pedicels

All of the populations had puberulent pedicels. Hitchcock

et al. (1959) describe Cj_ grandiflora flowers as being shorter

pediceled. According to Table XXV, this does not appear to be the

case.

e) Calyx

The presence or absence of puberulence on the calyx varies

between and within the populations and does not seem to be a

diagnostic character. But the length of the calyx compared to that

of the corolla was used as a diagnostic character in Newsom1s (1929)

monograph. She states that in C_» parviflora the calyx is "from half - 89 -

as long to almost equal the corolla length" and in C_» grandiflora the calyx is "1/3 - 1/2 the corolla length". In the populations studied, only two (Irvine's North, Elk Falls) had calyces which were less than

1/2 the corolla length. These are both very large-flowered populations but other large-flowered populations (e.g. Jack Point) did not show this characteristic. f) Corollas

In addition to corolla size C_, parviflora is said to differ from

C. grandiflora in the "possession of an almost erect and gibbous corolla tube in contrast to a declined and saccate tube" (Newsom, 1929).

The populations were studied for the angle of the corolla and whether the corolla was saccate or gibbous. Most of the populations examined had a corolla tube declined at about a 45° angle. Six pop• ulations had almost erect corollas but since these were found in both larger flowered- and small flowered-populations, there is no very good correlation between small flowers and erect corollas.

Three populations contained plants with corollas that were strongly

declined (about 90°)• Two of these were the very large-flowered

populations from Irvine's North and Elk Falls. The other was a

smaller-flowered population from Mill Hill II.

There is also no clear pattern in the distribution of saccate

and gibbous tubes. Many populations had both types of corolla tube

present, usually with their smaller flowers being gibbous and their

larger ones saccate. Again, the gibbous corolla tube does not

correlate with almost erect flowers. There does however seem to be a general trend toward gibbous corolla tubes in the smaller-flowered populations and saccate corolla tubes in the larger-flowered populations.

Another corolla character used by some authors to distinguish the two species is the relative size of the corolla limb versus the corolla tube (Piper, 1906; Henry, 1915; Peck, 1961), In C_, parviflora the corolla tube is said to be longer than the limb and in grandi flora the limb is not longer than the tube. In the populations studied, all were of the "parviflora" type in that the tube was longer than the limb, g) Flower size

There was a continuous range of flower sizes from 4 mm to 15 mm without any clear break. If the populations were to be divided into

two or more groups on the basis of flower size there would be con•

siderable overlap between the two classes no matter where the line

was drawn.

Discussion

Although there is considerable variation in vegetative charact•

eristics among and within the populations studied, there were no

correlations among the characters that would support dividing the

group into two species.

The floral characteristics are taxonomically more useful. There

is a general trend towards the association of saccate corolla tubes

with larger flowers and gibbous corolla tubes with smaller flowers. - 91 -

There is also a trend towards an association of declined (45°) to almost erect corolla tubes with smaller flowers and declined (45°) to strongly declined (90°) corolla tubes with larger flowers. How• ever, neither of these trends can be used to make a clear-cut division into two species.

The calyx character is more clear cut. In most populations, the calyx is greater than 1/2 the corolla length. In two of the largest-flowered populations, however, the calyx length is less than

1/2 of the corolla length. Using this character, it is possible to separate these two populations from the rest. These two populations also both have strongly declined corollas and saccate corolla tubes but these three characters are not entirely independent of one another. If calyces stay the same size but corollas are longer, the calyx will automatically be less than 1/2 the corolla length. And since the difference between saccate and gibbous is a matter of degree, if the corolla as a whole is smaller, it might be expected to have smaller gibbosity. The angle of the corolla will also

strongly influence the degree of gibbosity. These characters are

probably all different expressions of just one difference ~ smaller

versus larger corollas. Therefore, the separation of these two

populations from the rest is not sufficient to justify putting them

into a separate taxonomic species.

Therefore I propose to place all the entities now considered to

be C_ grandiflora Dougl, ex Lindl. and C&. parviflora Doug Vex Lindl.

into a single species; _C_. parviflora Dougl. ex Lindl. since this name

has published priority. In addition, I propose to erect two sub- 92 -

species, C, parviflora ssp, parviflora and C, parviflora ssp, grandiflora (Dougl,.ex Lindl,) Krause to express the morphological differences discussed above.

Although these two subspecies are somewhat arbitrary, they do recognize the two extremes in flower size that were formerly given species status. And they maintain the terms "parviflora" and

"grandiflora" to distinguish these extremes. - 93

Collinsia parviflora Dougl, ex Lindl, Bot, Reg, 13 : p, 1082 1827, ssp, parviflora

Anthirrhinum tenellum Pursh, Fl, Am, Sept. 421, 1814, Linaria tenella F, G, Dietr, Vollst, Lexik, Gaertn, Nachtr, 4:408, 1818, Collinsia tenella Piper. Contr, U.S. Nat. Herb, 11:496, 1906, Not C, tenella Benth. 1846. C. minima Nutt. Journ. Acad. Phila. 7:47, 1834. C. parviflora var. minima M. E. Jones, Contr. West. Bot. 12:69. 1908. C. grandiflora var. pusilla Gray, Syn. Fl. 21:256. 1878. C. pusilla Howell Fl. N.W. Am. 506. 1901. C. grandiflora ssp. pusilla Piper, Contr. U.S. Nat. Herb. 11:496. 1906. C. breviflora Suksd. W. Amer. Sci. 12:54. 1901. C. multiflora Howell, Fl. N.W. Amer. 506. 1901. C. diehlii M. E. Jones, Contrib. West. Bot. 12:68. 1908. C. parviflora var. diehlii Pennell in Abrams, 111. Fl. Pac. St. 3:778. 1951. C. parviflora forma alba English in St. John, Fl. S.E. Wash. 370. 1956. C. parviflora forma rosea Warren, Proc. Biol. Soc. Wash. 41:197. 1928. Stems ascending or erect, branched or unbranched, glabrate to puberulent, 2.4-40 cm tall; leaves various, serrulate to entire, ovate to lance-linear, obtuse to acute, labrate to puberulent, petiolate to occasionally sub-sessile, often purplish below, 0.6- 5 cm long, 0.3-1.9 cm wide, becoming smaller and bractiform in inflorescence; flowers in whorls, 2-7 at a node, often solitary below; pedicels puberulent to glandular-pubescent; calyx membranous below, puberulent to glabrate, scabrous margined, from 1/2 as long to almost equal the corolla length; calyx lobes subulate to linear-lanceolate, acuminate, ca, 1 mm wide; corolla usually declined (ca 45°) to almost erect, 4-10 mm in length, various shades of blue to purple, sometimes with white or whitish upper lip, rarely all white or magenta; fila• ments stout, glabrous; stigma 2-lobed; 3-5 mm long, slightly exceeded by calyx tips; seeds, usually 4-6, round-oblong, thick, smooth, reddish-brown or brown.

Collinsia parviflora Dougl. ex Lindl. ssp. grandiflora (Dougl. ex Lindl.) Krause. stat. n.

£i grandiflora Dougl. ex Lindl. Bot. Reg. 13:pi. 1107. 1827. C. grandiflora var. nana Gray, Proc. Am. Acad. 8:394. 1872. ! -94 -

Similar to parviflora ssp, parviflora; corollas 9-17 mm long; strongly declined (ca 90°); saccate corolla tube; calyx less than 1/2 the corolla length.

By placing the entire group into one species with two subspecies, both biological relationships and morphological differences are ex•

pressed. The single species expresses the interfertility as well as

the continuous nature of the variation within the group. But the

recognition of two subspecies, though somewhat artificial, recognizes

the fact that in spite of the seemingly continuous variation, the two

extremes are very different. - 95 -

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Anderson, J.P. 1961. Flora of Alaska and adjacent parts of Canada. Iowa State University Press, Amer. Iowa.

Bopp, M. 1959* Uber die bildung von anthocyan und leucoanthocyan an wundrandern. Zeitschrift fur Botanik 197-215*

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Davis, Ray J. 1952. Flora of Idaho. Wm. C. Brown Co. Dubuque, Iowa.

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Gorsic, J. 1957* The genus Collinsia V. Genetic studies in £4 heterophylla. Botanical Gazette 118(3): 208-223. Grant, V. 1963. The origin of adaptations. Columbia University Press, New York.

. 1971. Plant speciation. Columbia University Press, New York. - 96 -

Griffiths, A.J.F., Krause, G. and Ganders, F.R. 1977. A leaf spot polymorphism in Collinsia grandiflora (Scrophulariaceae). Canadian Jour* of Botany _5_5: 654-661.

Harborne, J.B. 1967. Comparative biochemistry of the flavenoids. Academic Press, London.

Hayhome, Barbara A. and Garber, E.D. 1968. The genus Collinsia XXIX. Preferential pairing in diploid, triploid and tetraploid interspecific hybrids involving stricta x Cj, concolor and related species. Cytologia 33(2): 246-255. Henry, J.K. 1915* Flora of southern British Columbia and Vancouver Island. W.J. Gage & Co. Ltd., Toronto.

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