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This is to certify that the

dissertation entitled
FLORAL EXPRESSION PATTERNS AND
GENETICS 0F DAY-NEUTRALITY IN OCTOPLOID FRAGARIA

presented by
Sedat Serce

has been accepted towards fulfillment
of the requirements for

Ph-D- degree infljnIMdjng and Genetic

 

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Date 10/08/2002

 

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6/01 cJCIRC/DateDuepss-sz

FLORAL EXPRESSION PATTERNS AND GENETICS OF DAY-NEUTRALITY IN
OCTOPLOID F RAGARIA

By

Sedat Serce

A DISSERTATION

Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of

DOCTOR OF PHILOSOPHY
Plant Breeding and Genetics Program

2002

ABSTRACT

FLORAL EXPRESSION PATTERNS AND GENETICS OF DAY-NEUTRALITY IN
OCTOPLOID FRAGARIA

By

Sedat Serge

The temperature and photoperiod interactions of a number of elite genotypes of F.
virginiana, F. xananassa, and F. chiloensis were studied in a series of growth chamber
experiments involving: 1) the critical day length of short day (SD) genotypes under 8, 9, 10,
and 11 h days at 18 °C; 2) the photoperiod characteristics of day-neutral (DN) and long day
(LD) types under 8 and 16 h days at 18 °C, and 3) the effect of temperature on flower bud
formation in DN genotypes under 12 h days at 18, 22, 26, and 30 °C. It was found that SD
plants were much less sensitive to photoperiod change than previously reported. Two elite
native DN genotypes LH 50—4 and RH 30 produced significantly more runners under LD and
cool conditions than the DN cultivars now grown, but were not heat tolerant.

Several different greenhouse and field methods were compared to score DN in
segregating populations. Scoring DN progeny within 100 days from germination was a poor
predictor of field performance. However, greenhouse screens were accurate in predicting
field performance, if the flowering behavior of individuals was followed through a whole
season. The percentage of DN progeny observed in our second year greenhouse results were

highly correlated with the subsequent field evaluations, and the families with the highest

flowering strength in the field also had the highest percentage of DNs in both greenhouse and
field screens.

To elucidate the inheritance of day-neutrality in octoploid F ragaria, crosses were made
between DN x SD and DN x DN types using F ragaria xananassa cultivars and elite clones
of F ragaria virginiana. Wide ranges in the percent of day-neutral progeny were found in the
various families (30 - 87% in DN x SD and 22 - 93% in DN x DN crosses), suggesting that
day-neutrality in octoploid strawberries is a quantitative trait and not regulated by a single,
dominant gene as previously suggested. Several other observations supported this conclusion
including: 1) Less than half the families produced 1:1 or 3:1 ratios of day-neutrals. 2) DN F.
virginiana genotypes produced significantly different percentages of DN progeny than DN F.
xananassa cultivars. 3) Two different DN parents crossed to the same short day genotype
produced different percentages of DN progeny. 4) None of the DN parents produced 100%
DN progeny, which would be expected homozygous dominant DN individuals existed. 5)
Some of the day-neutrality sources were more powerful than others in producing of day-
neutral progeny (e.g., 'Tribute' > 'Aromas' and RH 23 > Frederick 9). 6) Both general and

specific combining abilities for DN were significant.

I dedicate this work to the people of Yellibel Koyii

iv

ACKNOWLEDGEMENTS

I would like to thank my major professor Dr. James F. Hancock for his great
slippom instruction, and guidance throughout my doctorate studies. I am also grateful for
what I learned from the other studies in his laboratory, and his patience and enormous
editorial help during the writing process of this dissertation.

I would like to thank my committee members Dr. James Flore, Dr. Amy Iezzoni,
and Dr. James Kelly for serving on my committee and their extremely valuable
comments.

I would also like to thank the members of Dr. Hancock laboratory especially Pete
Callow for his help, support, and discussion.

I would like to acknowledge the support of all my great friends including Onur
Baser (Bilyfik Insan), Hfiseyin Yfice, Aydin Yfiksel, and Burak Yuran.

I express my appreciation to the Turkish Ministry of Education for their financial
support which made my graduate studies and priceless experiences in United States
possible.

Gratitude is also due to my parents, Nadiye and Hfiseyin Serge, and my sisters and
brothers for their love and support. Finally, I thank Seckin Serce for always supporting

me and believing in my abilities.

TABLE OF CONTENTS

LIST OF TABLES ........................................................................................................... viii
LIST OF FIGURES ......................................................................................................... ....x
ABBREVIATIONS .......................................................................................................... xii
CHAPTER 1
TEMPERATURE AND PHOTOPERIOD REGULATION OF F LOWERING IN
OCTOPLOID FRAGARIA
Introduction ................................................................................... 1
EXperiment 1 ................................................................................. 8
Material and Methods .................................................................................. .8
Results and Discussion .............................................................................. .9
Flower numbers ................................................................. 9
Runner numbers ............................................................... 11
Dry weights ..................................................................... 12
Experiment 2 ................................................................................ 19
Material and Methods .................................................................................. 19
Results and Discussion .............................................................................. 20
Flowering response to LD and SD .......................................... 20
Runnering response to LD and SD ......................................... 21
Response of dry weights to LD and SD .................................... 22
Experiment 3 ................................................................................ 26
Material and Methods .................................................................................. 26
Results and Discussion .............................................................................. 27
CHAPTER 2

INHERITANCE OF DAY-NEUTRALITY TN FRAGARIA xANANASSA AND
FRAGARIA VIRGINIANA

Introduction .................................................................................. 38
Experiment 1 ................................................................................ 43
Material and Methods .................................................................................. 43
Results and Discussion .............................................................................. 47
Day-neutrality evaluation methods ......................................... 47

vi

Horticultural traits ............................................................. 48

Conclusions .................................................................... 49

Experiment 2 ................................................................................ 59
Material and Methods .................................................................................. 59

Results and Discussion .............................................................................. 61

Chi-square analysis ........................................................... 61

General and Specific Combining Abilities ................................. 62

Conclusions .................................................................... 63

REFERENCES .................................................................................................................. 71

vii

LIST OF TABLES

Table 1. Analysis of variance for flower and runner numbers, and dry weights of
strawberry genotypes grown at 18 °C and either 8, 9, 10, or 11 h days ................... 13

Table 2. Means and standard errors (in parenthesis) for flower and runner numbers, and
dry weights of strawberry genotypes grown at 18 °C and either 8, 9, 10, or 11 h
days ........................................................................................................................ .14

Table 3. Regression lines, R2, and significance (P) for flower and runner number, and
dry weights of strawberry genotypes grown at 18 °C and either 8, 9, 10, or 11 h
days .......................................................................................................................... 15

Table 4. Means and standard errors for flower and runner numbers, and dry weights of
strawberry genotypes grown at 18 °C and either 8 or 16 h days .............................. 23

Table 5. Analysis of variance for flower and runner numbers, and dry weights of
strawberry genotypes grown in growth chambers at 12 h days and 18, 22, 26, and
30 °C ......................................................................................................................... 31

Table 6. Means and standard errors (in parenthesis) for flower and runner numbers, and
dry weights of strawberry genotypes grown in growth chambers at 12 h days and
18, 22, 26, and 30 °C ................................................................................................ 32

Table 7. Regression lines, R2, and significance (P) for flower and runner number, and
dry weights of strawberry genotypes grown in growth chambers at 12 h days and
18, 22, 26, and 30 °C ................................................................................................ 34

Table 8. The genotypes crossed in a partial-diallel fashion to study the interaction of
different evaluation method of day-neutrality ......................................................... 45

Table 9. Parentage of day-neutral progeny in strawberry families grown in a greenhouse
at Michigan State University, East Lansing, Mich., and in the field at the Southwest
Michigan Research and Extension Center Benton Harbor, Mich. in 2000 and
2001 ................................................................................................. 51

Table 10. Correlation coefficients, significance, and number of individuals in strawberry
families grown in a greenhouse at Michigan State University, East Lansing, Mich.,
and in the field at the Southwest Michigan Research and Extension Center Benton
Harbor, Mich. in 2000 and 2001 ............................................................. 53

Table 11. Mean and standard deviations for horticulturally important traits in strawberry
families grown in a greenhouse at Michigan State University, East Lansing, Mich,

viii

and in the field at the Southwest Michigan Research and Extension Center Benton

Table 12. Correlation between percentage of day-neutral progeny and several
horticulturally important traits in strawberry families grown in a greenhouse at
Michigan State University, East Lansing, Nfich., and in the field at the Southwest
Michigan Research and Extension Center Benton Harbor, Mich. in 2000 and
2001 ............................................................................................. 58

Table 13. The family numbers of genotypes crossed in a partial-diallel fashion to study
the genetic of day-neutrality in strawberries .............................................. 60

Table 14. Proportion of short day (SD) and day-neutral (DN) progeny generated in SD x
DN crosses. Chi-square tests were made assuming DN was regulated by a single
dominant gene (1 :1). The families were grown in a completely randomized design
at the Michigan State University Horticulture Farm, East Lansing, Mich. in
2002 ............................................................................................. 65

Table 15. Proportion of short day (SD) and day-neutral (DN) progeny generated in DN x
DN crosses. Chi-square tests were made assuming DN was regulated by a single
dominant gene (3:1). The families were grown in a completely randomized design
at the Michigan State University Horticulture Farm, East Lansing, Mich. in
2002 ............................................................................................. 67

Table 16. General and specific combining ability (GCA and SCA) for percent day-
neutral progeny in F. xananassa x F. virginiana families grown in a completely
randomized design at the Michigan State University Horticulture Farm, East
Lansing, Mich. in 2002 ...................................................................... 69

ix

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LIST OF FIGURES

Figure 1. Mean flower number of strawberry genotypes grown at 18 °C and either 8, 9,
10, or 11 h days. The bars represent standard errors ............................................... 16

Figure 2. Mean runner number of strawberry genotypes grown at 18 °C and either 8, 9,
10, or 11 h days. The bars represent standard errors ............................................ 17

Figure 3. Mean dry weight of strawberry genotypes grown at 18 °C and either 8, 9, 10, or
11 h days. The bars represent standard errors ......................................................... 18

Figure 4. Mean total flower number of strawberry genotypes grown at 18 °C and either 8
or 16 h days. The bars represent standard errors .................................................. 24

Figure 5. Mean total flower number of strawberry genotypes grown at 18 °C and either 8
or 16 h days. The bars represent standard errors ........................................ 25

Figure 6. Mean flower number of strawberry genotypes grown in growth chambers at 12
h days and 18, 22, 26, and 30 °C. The bars represent standard errors .................... 35

Figure 7. Mean runner number of strawberry genotypes grown in growth chambers at 12
h days and 18, 22, 26, and 30 °C. The bars represent standard errors .................... 36

Figure 8. Mean dry weight of strawberry genotypes grown in growth chambers at 12 h
days and 18, 22, 26, and 30 °C. The bars represent standard errors ........................ 37

Figure 9. Association of several day-neutrality scoring methods in strawberry families
grown in a greenhouse at Michigan State University, East Lansing, Mich., and in
the field at the Southwest Michigan Research and Extension Center Benton Harbor,
Mich. (DNl-F lowering within 100 days from germination in a greenhouse in 2000;
DN2-F lowering before 9/9 in field in 2000; DN3-Flowering under both short and
long days in a greenhouse in 2001; DN4-Flowering under both short and long days
in field in 2001) ................................................................................................... 54

Figure 10. Association of several day-neutrality scoring methods in strawberry families
grown in a greenhouse at Michigan State University, East Lansing, Mich., and in
the field at the Southwest Michigan Research and Extension Center Benton Harbor,
Mich. (DN4-Flowering under both short and long days in field in 2001; DNS-
Flowering on their newly-formed runner in field in 2001; Flowering strength ratio
of 0-10 (10 having the most flowers during the second cycle of flowering) in field

Figure 11. Percentage of day-neutral progeny in strawberry families of DN x SD crosses
grown in a completely randomized design at the Michigan State University
Horticulture Farm, East Lansing, Mich. in 2002. The white column represents F.
xananassa x F. xananassa, while gray and black columns represent F. xananassa x
F. virginiana and F. virginiana x F. virginiana crosses. The line represents the
expected percentage assuming day-neutrality was regulated by a single dominant
gene ............................................................................................... 66

Figure 12. Percentage day-neutral progeny in strawberry families of DN x DN crosses
grown in a completely randomized design at the Michigan State University
Horticulture Farm, East Lansing, Mich. in 2002. The white column represents F.
xananassa x F xananassa, while gray and black columns represent F xananassa x
F. virginiana and F. virginiana x F. virginiana crosses. The line represents the
expected percentage assuming day-neutrality was regulated by a single dominant
gene ............................................................................................. 68

Figure 13. Percentage of day-neutral progeny in strawberry families grown in a
completely randomized design at the Michigan State University Horticulture Farm,
East Lansing, Mich. in 2002. The white column represents F. xananassa cultivars,
while black columns represent F. virginiana genotypes. ‘ The error represent
standard errors ................................................................................. 70

xi

°C
ANOVA
CDL

DN

DN 1
DN2
DN3
DN4
DNS
FSR

LD
MSU
PAR
QTL
SD
SE

ABBREVIATIONS

Degree Celsius

Analysis of variance

Critical day length

DaY(S)

Day-neutral or day-neutrality
100 days to flowering

1St year field flowering

2nd year greenhouse flowering
2"(1 field flowering

Runner flowering

Flowering strength rating
Gram

Hour(s)

Long day

Michigan State University
photosynthetically available radiation
Quantitative trait locus/loci
short day

Standard error

slope of the regression line
value observed (in regression equation)

xii

CHAPTER 1
TEMPERATURE AND PHOTOPERIOD REGULATION OF F LOWERING IN

OCTOPLOID FRAGARIA

Introduction

All three photoperiodic types, short day (SD), long day (LD), and day-neutral (DN)
exist in strawberries F ragaria sp., although most of the octoploid commercial varieties
now grown are either SD or DN (Hancock, 1999). Temperature and photoperiod
interactions play a very important role in the flowering of strawberries (Darrow 1936;
Dumer et al., 1984). DN strawberry cultivars do not flower at high temperature (< 28 °C)
(Durner et al., 1984). SD cultivars are facultative, meaning they form flower buds under
SD conditions when temperatures are moderate, but they can form flowers under non-
inductive LD conditions when temperatures are below 15 °C (Guttridge, 1985; Darrow
1936; Larson 1994). Temperature also significantly affects the number of inductive
cycles (days) and the critical day length (CDL) of SD strawberry genotypes (Guttridge,
1985)

Critical day length is associated with how early cultivars begin forming flower buds
in the summer and this may affect yield. For example, a cultivar with ~12 h CDL will
start initiating flower buds in September in Michigan, while another one with ~11 h CDL
would start in October. The cultivar initiating flower buds earliest would have more time

to produce flower promordia before the onset of the first severe frost of the fall.

While numerous sources of day-neutrality have been identified in strawberries over
the years, the first successful introgression into commercial octoploid strawberries was
done by Bringhurst and Voth (1984) at the University of California, Davis. They
transferred genes from a native clone of F ragaria virginiana ssp. glauca from the
Wasatch Mountains of Utah and were able to generate commercially useful genotypes
within a few generations of backcrossing into F ragaria xananassa. Currently, DN
cultivars are grown on about 60% of the California production area, but remain a minor
component of strawberry production in the eastern US. (Hancock, 1999).

The same source of day-neutrality has been used to produce new DN cultivars in
many North American breeding programs (Sjulin and Dale, 1989; Hancock et al., 1990).
However, while the Wasatch source for day-neutrality has proVen useful in California
and other Mediterranean climates, it has not performed well in other parts of North
America (Dale et al., 2002). Cultivars that have the Wasatch source suffer from summer
heat in continental climates. In fact, flower bud initiation is completely inhibited at 30/26
°C day/night temperatures (Galletta et al., 1981; Durner et al., 1984; Strick 1985). The
DN cultivars have, at best, reduced yields and small, soft fruits in the middle of the
summer (Draper et al., 1981). In addition, while early reports indicated that day-
neutrality performed as a single dominant gene in the California gene pool (Bringhurst
and Voth, 1978; Ahmadi et al., 1988), it does not appear to act in a similar fashion in
genetic backgrounds elsewhere, and the D "gene" varies in expressivity (Barritt et al.,
1982; Nicoll and Galletta, 1987; Hancock et al., 2002).

In a number of recent germplasm surveys, several elite clones of F virginiana have

been selected (Frederick 9, LH 50—4, and RH 30) that may be DN as they were multiple

cropping in the field and proved useful in breeding multiple cropping progeny (Hancock
et al., 2001 a and b; Serge and Hancock, 2002; Serge et al., 2002). However, the
photoperiod relationships of these elite genotypes have not been tested.

Temperature and photoperiod are the most important environmental factors that
regulate the transition from vegetative to floral growth in strawberries (Darrow 1936;
Dumer et al., 1984). Although the transition fi'om vegetative to floral growth is a
continuous process, it can be divided into three stages; induction, initiation, and
differentiation (Dumer and Poling, 1985). Floral induction takes place in the leaves after
the appropriate photoperiod and/or temperatures exposure. The physiological and
morphological changes that occur in meristems after the perception of these stimuli in the
leaves is called initiation. The development of specific floral organs or of flowers on a
single inflorescence is referred to as differentiation (Dumer and Poling, 1988). Visual
expansion of flowers is called flower development.

Investigators have used a number of different terminologies to describe photoperiod
and temperature regulation of flowering sensitivity in strawberries including short day or
Junebearing vs. everbearing (Clark, 1937; Powers 1954), as well as perpetual vs. non-
perpetual (Richardson, 1917). Strawberries have also been called single, double, or
multiple cropping based on how many times they flower each year. In North America
and Europe, flower bud initiation takes place in late September or early October in
Junebearering and SD types (Goff, 1903; Guttridge, 1958; Jahn and Dana, 1969),
regardless of plant age (Hill and Davis, 1929; Schilletter and Richey, 1930; Schilletter
and Richey, 1931). In everbearing and perpetual flowering cultivars, flower bud

initiation occurs at least twice during the season during long days (Darrow and Waldo,

1934). Day-neutral cultivars were introduced in the late 19705 and they initiate flower
buds cyclically all season as long as temperatures are below 26 °C.

Sudds (1927) was the first investigator to suggest that flowering was regulated by
photoperiod, when he obtained accelerated floral initiation in plants of 'Howard 17' by
maintaining them under 8 h day length. The early studies of Darrow and Waldo (1934)
resulted in the classification of strawberries as Junebearer and everbearer. They
concluded that the flower bud formation occurs in Junebearer types under short day
conditions (less than 14 h), and under long day conditions (more than 12 h) in the
everbearer ones. Darrow (1936) also studied the temperature effect on strawberry
development and found that high temperatures (21 °C) favored runner formation.
Hartrnann (1947 a and b) studied photoperiod and temperature effects on flower
development in several cultivars ('Blakemore', 'Fairfax', 'Marshall', and 'Missionary'). He
followed the response of the cultivars to 10 or 15 h days at 16 and 21 °C temperatures,
after acclirnating the plants under long days (15 h). He found that at 16 °C all cultivars
flowered regardless of day length treatments. He also found that none of the cultivars
flowered under 15 h days and 21 °C treatments, while only ’Fairfax' flowered under 10 h
days and 21 °C temperature. His results supported the earlier conclusions of Darrow and
Waldo (1934) and Darrow (1936).

Dumer et al. (1984) studied the interaction between photoperiod x temperature
using representatives of Junebearing, everbearing, and day—neutral types. They held
plants at 21 °C and examined their flowering response to 9 h SD, 16 h LD, and 9 h SD
with a 3 h night interruption. They found that the flowering of DNs is little affected by

photoperiod. SD plants had highest inflorescence numbers under SD (SD = 7.5 and LD =

0.8), while LD had highest inflorescence numbers under LD plants had the (SD = 1.5 and
LD = 5.0). They also studied the effect of four day/night temperature treatments, the

l8/ 14, 22/18, 26/22, 30/26 °C. Under SD conditions, all types flowered only in 18/14 and
22/18 °C treatments, and not at 26/22, 30/26 °C. The mean inflorescence numbers at
22/18 °C was 0.3, 0.5, and 1.3 for SD, LD, and DN types, respectively. Based on these
results they ranked the sensitivity to high temperature as SD > LD > DN.

Reports indicated that from 7 -24 days was the minimum number of photo inductive
cycles necessary to induce flowering in short day plants, depending on temperature
treatments (Hartrnann, 1947 a and b; Went, 1957; Guttridge, 1985; Larson, 1994). Ito
and Saito (1962) studied the temperature effect on photo inductive cycles and found that
longer photo induction periods are required at higher temperatures. Only 10 cycles were
needed for floral induction under 8 h photoperiods, at 30 °C more than 20 cycles were
necessary. Under 16 h photoperiods, 10 cycles were needed for floral induction at 9 °C,
whereas 16 cycles were required at 17 °C.

To express their full potential, SD cultivars often require a chilling period (<7 °C),
although the cultivars developed for warm regions do not always need a chilling period
(Darrow, 1933). The ability to grow well in North American greenhouses during the
short days of October, November and December was used as an indicator of a cultivar's
regional adaptation (Amey, 1954). The cultivars well-adapted to cooler regions generally
do not grow well during this period and enter a rest period, while those adapted to
warmer climates continue to grow (Darrow and Waldo, 1934).

A new type of strawberry was recently developed in Israel for tropical and

subtropical environments (Izsak and Izhar, 1984; Izhar and Izsak, 1995). This type of

cultivar, called infia short day, can initiate flower bud under longer light regimes (13.5 to
14 h) and higher temperatures (10 - 26 °C) than traditional short day plants, and they do
not have a chilling requirement.

While the distinction between SD, LD, and DN types was elegantly demonstrated
by Dumer et al. (1984), several other important studies which have provided key
information on the regulation of flower development in the three photoperiod types
(Nicoll and Galletta, 1987; Yanagi and Oda, 1989). These studies have lead investigators
to believe that the photoperiod response is continuous rather than discontinuous. Darrow
(1966) suggested that strawberry genotypes actually range continuously from obligate
short day to facultative short day to complete day-neutrals, and indicated that DN types
vary in their flowering expression from weak to strong. The infra short day types are
thought to be in the middle of this range (Izsak and Izhar, 1984). Nicoll and Galletta
(1987) tested the temperature and photoperiod response of strong, weak, and very weak
day-neutrals, as well as older LD and SD types using Dumer's maximum temperature
regimes. The DN types were ranked based on the proportion of their daughter plants that
flowered during the summer. Their results were in agreement with Darrow (1966), as
they observed a continuous responses in flowering to photoperiod: all the plants flowered
once, but, none of the SD types re-flowered, 27% of very weak day-neutral re-flowered,
and 100% of the DNs and LD re-flowered. In addition, there were significant differences
among weak and strong DN and LD types in their intensity of flowering, fruit set, fruit
number and fi'uit weights.

Runnering is stimulated by long days and high temperatures in strawberries.

Darrow (1936) and Durner er al. (1984) demonstrated that if a clone produces any

runners, it will do so under long days and higher temperatures. In a greenhouse study,
Darrow tested the runnering ability of several cultivars grown in September with a 16 h
day at 21.1 °C, 15.5 °C, or 12.8 °C temperatures. Among these temperature treatments,
only 21.1 °C temperature yielded runners. Dumer et al. (1984) studied the effect of
temperature and photoperiod using combination of long day, short day, and night
interruptions and 16, 20, 24, and 28 °C temperature treatments. On average, 2.0, 1.2, and
0.4 runners per plant were observed for long day, night interruption, and short day
photoperiods, respectively. While there were no runners produced in SD at temperatures
below 24 °C, LDs and DNs generated similar numbers of runners under short days
regardless of temperature treatments. Under long days, however, runners were observed
in all the temperature treatments.

Smeets (1980) studied runner formation in 'Revada' and 'Rabunda' held under
combinations of 14, 20, and 26 °C for 8, 16, 24 h days. Although runners were observed
in all treatments, 20 and 26 °C treatments generated significantly more runners than 14
°C, while 16 and 24 h days generated significantly more runners than the 8 h day length
treatment. Sonsteby (1997) also found higher rtmner numbers under high temperatures in
an experiment where he studied the effect of 9, 15, and 21 °C temperature regimes at 8
and 24 h days using 'Korona', 'Elsanta', 'Bounty’, and 'Senga Sengana'.

In this chapter, temperature x photoperiod interactions of a number elite genotypes
were studied in a series of growth chamber experiments involving: 1) the CDL of SD
genotypes under 8, 9, 10, and 11 h days at 18 °C, 2) the photoperiod characteristics of
DN and LD types under 8 and 16 h days at 18 °C, and 3) the effect of temperature on

flower bud formation in DN genotypes under 12 h days and 18, 22, 26, and 30 °C.

EXPERIMENT 1

Material and Methods

This experiment was conducted to determine the required CDL and period of
induction for a wide range of putative SD genotypes. The representatives and their
region of origin were the F. xananassa cultivars 'Allstar' (Mid-Atlantic), 'Chandler'
(California), and 'Honeoye' (New York), F. chiloensis F RA 0024 (central Chile), FRA
0368 (Alaska), and F. virginiana Eagle 14 (Ontario).

Runners from each genotype were gathered fiom a field planting at the Michigan
State University (MSU) Horticultural Research Farm (East Lansing, Mich.) on 8/30/01
and placed under mist for rooting. On 9/10/01, 40 rooted runners were potted into 14 x
12 x 12 cm pots and set in a greenhouse where they were maintained under 13 h day
lengths using supplementary lights (~8OO umol s'l m'z). On 10/31/01, 40 plants of each
genotype were transferred into four growth chambers at 18 °C for 8, 9, 10, or 11 h days.
In each chamber, photosynthetically available radiation (PAR) was initially varied with
day length so that the total energy received was equal (~800, 710, 640, and 580 mol s'l
m'2 for 8, 9, 10, and 11 days, respectively). After 15 and 30 days (on 11/14/01 and
11/30/01), 5 plants of each genotype fi'om each chamber were placed in a greenhouse
held at 13 h day length using supplementary lights (~800 pmol s'l m‘z). The number of

flowers and runners produced by each plant were recorded on 11/15/01, 11/25/01, and

12/06/01. Total dry weights were also determined for each plant after washing their roots
free of soil and holding them at 72 °C for 3 days.

The initial analysis indicated that both 15 and 30 days induced the same number of
flowers and runners. As a result, these two treatments could be treated as different blocks
in evaluating the effect of the different day lengths. Analysis of Variance (ANOVA)
tables, means, and standard errors (SEs) were calculated for all variables using the SAS
program (SAS Institute, Cary, N.C.). Regression lines were fitted to the average values
of each genotype at each day length to determine the trend of the genotype’s performance

across day length.

Results and Discussion

Flower number: The number of flowers formed by all genotypes was not
significantly different under either the 15 or 30 day induction period (Table 1). The
observations indicate that at cool temperatures, such as 18 °C, the number of induction
cycles required to form flower buds is less than 15 days for the genotypes used in this
experiment. Although CDL has been reported to be between 7-24 depending on
temperature (Hartrnann, 1947 a and b; Went 1957; Larson, 1994), 7-14 cycles is
generally considered adequate under cool temperatures (Guttridge, 1985). Thus, our
results are in agreement with the literature.

All genotypes produced one inflorescence, regardless of photoperiod. Varying
numbers of flowers were observed depending on photoperiod; however, there was not a

consistent trend (Table 2 and Figure 1). The average number of flowers across all

genotypes for 8, 9, 10, and 11h days were 4.2, 3.5, 3.5, and 5.1, respectively (Table 2).
Likewise, the individual genotypes produced different numbers of flowers under different
photoperiods, but there were few consistent trends (Table 3). Eagle-14 had more flowers
under LD than SD and showed a positive increase as day length was increased (y = - 6.8
+ 1.3 x), while 'Honeoye' showed an overall decline in flower numbers as day length was
increased (y = 9.9 - 0.7 x) (Table 3). However, all the other genotypes displayed little
relationship between flower numbers and day length, with slopes less than b = 0.2 (Table
X). None of the regression lines for individual genotypes were significant (P < 0.05).

The flowering trend of 'Honeoye' was not significant, but it did fit the typical,
quantitative SD model reported by others (Darrow 1936; Darrow and Waldo, 1934;
Dumer et al., 1984). The flat regression lines of 'Allstar', 'Chandler', FRA 0024, and
F RA0368 indicates that day length between 8 - 11 h days does not have a significant
differential effect on flower bud initiation at 18 °C. Such insensitivity to day length has
not been reported previously in SD plants.

The positive relationship between photoperiod and flower production in Eagle-l4,
while also not significant, is at least suggestive that it is DN or LD rather than SD. In
previous studies, Eagle-l4 has not flowered in the late summer or fall in the field as
would be expected for a LD or DN genotype (Hancock et al., 2001 a), but high summer
temperatures may have inhibited its flower production (Dumer et al., 1984; Hartnrann,
1947 a and b). Occasional flowers have been observed on Eagle-l4 during long summer
days in our cooled greenhouse (Callow, personal communication), and Eagle-14 has
produced a high number of DN progeny in crossing studies (Hancock et al., 2002;

Chapter 2).

10

It is not known why our flowering response patterns were so different from the
literature, but all previous studies involved F. xananassa cultivars that are no longer
grown. Perhaps, selection for high yields has altered the response pattern of some of our
modern cultivars. A broader comparison of native germplasm might be warranted to
uncover the variability available for photoperiod response in strawberry.

Runner number: The day length treatment, genotypes, and their interaction were
highly significant for runner number (Table 1). However, 'Allstar' and 'Honeoye' did not
runner at all, while 'Chandler' (mean = 0.1) and FRA 0024 (mean = 0.3) produced only
negligible runner numbers (Table 2 and Figure 1). Only FRA 0368 and Eagle-14
generated significant numbers of runners in any treatment. The average runner numbers
of FRA 0368 were 0.7, 0.6, 0.4, and 2.9 (mean = 0.9), while Eagle-l4 had 2.0, 3.0, 2.6,
and 3.1 for 8, 9, 10, and 11h days, respectively (mean = 2.7) (Table 2). The slopes of the
regression lines (b = 0.3 for Eagle-l4 and b = 0.8 for F RA 0368) revealed a progressive
trend as day length advanced, although these regression lines were not significant (Table
3).

The positive relationship observed between photoperiod and runner numbers is
consistent with the literature, as strawberries has been reported to have more runners
under LD (Darrow 1936; Darrow and Waldo 1934; Dumer et al., 1984; Larson, 1984).
Runner numbers have also been reported to be highest under hot temperatures (Dumer et
al., 1984; Heide, 1977, Smeets, 1980; Sonsteby, 1997). This may be why many of the
genotypes produced few runners overall as our induction temperatures were cool (18 °C).

Dry weights: The main effect of day length treatment was not significant for dry

weights (Table 3 and Figure 3) suggesting that most of the differences observed in flower

ll

and runner numbers were not caused by PAR, but photoperiod. 'Allstar' and 'Chandler'
did have slightly positive (y = 12.8 + 1.1 x ) and negative (y = 16.6 - 0.8 x) regression

lines, but these were not significant.

12

Table 1. Analysis of variance for flower and runner numbers, and dry weights of
strawberry genotypes grown at 18 °C and either 8, 9, 10, or 11 h days.

 

 

 

Source df Flower no. Runner no. Dry weight (g)
Block 1 32.3 0.2 4.6

Day length (D) 3 34.5“ 2.0" 5.2
Whole-plot error 35 8.7 0.4* 5.5
Genotype (G) 5 103.2" 42.2" 569.5”
D*G 15 15.2* 1.3" 15.4"
Error 180 7.8 0.3 6.8

 

*, " Significant at 0.05 and 0.01, respectively.

13

 

 

 

 

14

 

 

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15

Figure 1. Mean flower number of strawberry genotypes grown at 18 °C and either 8, 9,
10, or 11 h days. The bars represent standard errors.

* I911
10 _. ITOh
I11h

 

Flower number
0)

 

 

 

 

 

 

FRA 0024
FRA 0368
'Chandler'
'Honeoye'

16

Figure 2. Mean runner number of strawberry genotypes grown at 18 °C and either 8, 9,

10, or 11 h days. The bars represent standard errors.

 

Runner number

41

 

 

Eagle-14 "

FRA0024 '

FRA 0368

 

17

 

’Allstar'

'Chandler’

08h
39h
I10h
I11h

 

'Honeoye'

Figure 3. Mean dry weight of strawberry genotypes grown at 18 °C and either 8, 9, 10, or
11 h days. The bars represent standard errors.

   

20 1 08h

a 9n

. l10h

16 4. I11h
12 J

Dry weight (9)

 

 

 

 

 

FRA 0024
FRA 0368
'Chandler‘
'Honeoye'

18

EXPERIMENT 2

Material and Methods

This experiment was designed to determine the photoperiod sensitivity of a number
of elite wild clones and to verify whether the old “everbearer” strawberry cultivars are
DN or LD. Frigo plants were used of the everbearing cultivars 'Fort Laramie' and
'Quinalt'; DN cultivars 'Aromas' and 'Tribute', putative DN F. virginiana selections
Frederick 9, LH 50-4, RH 30; SD F. chiloensis selection F RA 0368. Further information
on the elite wild clones is available in Hancock et al. (2001 a and b) and
www.berrygenetics.com. The cultivars were purchased from Gurney’s Seed & Nursery
(Y ankton, SD.) and the dormant F. virginiana clones were dug from MSU Research
Farm, East Lansing, in April 2001.

Ten plants of each genotype were potted into 14 x 12 x 12 cm pots with a planting
mix purchased from the Michigan Peat Company (Houston, TX). The potted plants were
placed in a completely randomized design in a greenhouse at MSU on 4/11/2001 and
were held for 3 months under 12 h day length maintained with supplementary light (~800
umol s'1 m'z). During this period, all of the genotypes flowered. On 7/11/2001 the plants
were randomly placed into two growth chambers at 18 °C, 8 h day length and 800 mol
s" m'2 PAR or 18 °C, 16 h day lengths with 400 pmol s'1 m'2 PAR. PAR was varied with
day length so that the total energy received by the plants in each chamber was equal. The

plants were given a 7-week induction period, and then flower and runner numbers were

19

recorded on 9/4/01, 9/10/01, 9/17/01, and 9/22/01. No data were recorded on the flowers
and runners that developed before this time, as they could have been initiated before the
plants were placed in the greenhouse. On 9/25/01, each plant was dried at ~72 °C for 3
days and weighed. Means and SEs were calculated for all variables using the SAS
program (SAS Institute, Cary, NC). Because the growth chamber conditions were not
replicated, ANOVA tables could not be constructed; however, SEs could be calculated as

the genotype means represented 5 plants within each treatment.

Results and Discussion

Flowering response to LDgnd SD: Two types of flowering patterns were observed:
1) flowering under both LD and SD conditions ('Aromas', 'Tribute', FRA 0368, Frederick
9, LH 504, RH 30) and 2) flowering under LD but not SD conditions ('Fort Laramie' and
'Quinalt') (Table 4 and Figure 4). These patterns represent what would be expected of
DN and LD types, respectively. The mean inflorescence number ranged from 1.0 (F RA
0368) to 3.0 (Frederick 9) in the SD types and 0.2 (RH 30) to 3.4 (’Tribute') in the LD
types. Frederick 9 had the highest total number of flowers (15.4) under SDs while
'Tribute' was the most productive genotype under LDs (25.6) (Table 4). 'Fort Laramie'
produced an average of 2.6 inflorescences and 13.4 total flowers, while 'Quinalt'
generated 3.4 inflorescences and 23.0 total flowers (Table 4 and Figure 4).

These results indicate that the multiple cropping behaviors of Frederick 9, LH 50
and RH 30 that were previously observed in the field were due to their being DN and not

LD plants. FRA 0368 also appears to be DN, even though it is not typically multiple

20

cropping in the field. Perhaps, high summer field temperatures have inhibited floral
production in FRA 0368, as it is from Alaska where such high temperatures are rare. We
did observe some flowers on F RA 0368 in the field in the relatively cool weather of
September 2001 (Osborn, personal communication). The fact that DN LH 50-4 and RH
30 produce many runners under long day and cool conditions is exciting, as one of the
limitations of modern DN cultivars is their inability to runner (Dale et al., 2002).

Runnering response to LD am] SD: 'Aromas', 'Tribute', Frederick 9, and 'Fort
Laramie' produced no runners under either LB or SDs (Table 4). F RA 0368 and 'Quinalt'
produced runners under just LDs (2.5 and 0.2), while LH 50-4 and RH 30 produced
runners under both SD and LD conditions. The mean runner numbers were 2.5 (F RA
0368) and 0.2 ('Quinalt') (Table 4). LH 50-4 generated 1.4 runners under SD and 4.4
under LDs, and RH 30 produced 0.2 under SD and 3.8 under LD (Table 4).

It is not known why some of the genotypes did not runner under LDs, as numerous
studies have reported that runnering in strawberries is a LD response (Darrow 1936;
Durner et al., 1984; Dumer et al., 1984; Smeets, 1980; Sonsteby 1997). Perhaps
runnering in 'Aromas', 'Fort Laramie', Frederick 9, and 'Tribute', is more sensitive to cool
temperature than the other genotypes. As previously mentioned in the Experiment 1,
warm temperatures are generally thought to be stimulatory to runner production than cool
but few modern cultivars have been examined. There may also have been differential
sensitivities among genotypes to photoperiod, as LH 50-4 and RH 30 produced runners
under both long and short days; while the rest of the genotypes produced runners only
under LD conditions or none at all. It has been assumed that the genotypes that were

photoperiod insensitive for flowering retained LD response of runnering (Dumer et al.,

21

1984; Heide, 1977; Dumer et al., 1984; Smeets, 1980; Sonsteby, 1997). LH 50-4 and RH
30 may be photoperiod insensitive for both of these developmental responses.

Respgnse of g weigl_rts to LD and SD: Overall mean dry weights were 23.1 g and
26. 6 g for SD and LD types, respectively (Table 4 and Figure 5). Some differences were
also observed among genotypes between the two day length treatments. For example,
FRA 0368 had higher mean dry weights (25.3 g) under LD than SD (22.9 g), whereas
'Quinalt' had higher mean dry weights (30.8 g) under SD than LD (22.0 g) (Table 4 and
Figure 5). These results indicate that the PAR adjustments made at the beginning of the
experiment for day length treatments may not have been maintained throughout the
experiment. This imbalance may have influenced relative numbers of flowers in the two
treatments, but the main goal of the experiment was to determine whether a genotype
flowered under LD or SD conditions. The modest differences in PAR were unlikely to

have impeded the overall flowering response of genotypes.

22

 

 

 

 

 

 

 

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23

Figure 4. Mean total flower number of strawberry genotypes grown at 18 °C and either 8
or 16 h days. The bars represent standard errors.

30 08h
I16h

N N
O 01
r _ _ do;

 

Flower number
A
0|

—L
o

     

 

 

 

FRA 0368
Frederick 9
LH 50-4
RH 30
'Aromas'
'Tribute'
'Qumalt‘

'Fort Laramie'

24

Figure 5. Mean total flower number of strawberry genotypes grown at 18 °C and either 8
or 16 h days. The bars represent standard errors.

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RH 30
'Aromas'
'Tnbute'
'Qumalt’

'Fort Laramie'

25

EXPERIMENT 3

Material and Methods

This experiment was designed to evaluate the effect of temperature on flowering in
a number of old everbearing cultivars developed in North Dakota where very warm
summer temperatures occur, and several elite F. virginiana genotypes collected fiom a
wide range of environments. The goal was to identify genotypes that can form flower
buds under higher temperatures (>26 °C). The genotypes studied were the elite F.
virginiana genotypes Brighton-3 (Utah), LH 30-4 (Montana), LH 39-15 (Montana), LH
40-4 (Montana), LH 50-4 (Montana), RH 23 (Minnesota), RH 30 (Minnesota), RH 43
(Alaska), RH 45 (Alberta); and F Xananassa 'Aromas', 'Fort Laramie', 'Ogallala', and
'Tribute'. Further information on the native elite genotypes can be found at Sakin et al.
(1997), Hancock et a1. (2001 a and b) and www.berrv2enetics.com. The cultivars were
purchased from Gurney’s Seed & Nursery (Yankton, SD.) and the dormant F. virginiana
clones were obtained from Michigan State University Horticultural Research Farm in
August 1999.

Twenty eight plants of each genotype were potted on 8/5/99 into 14 x 12 x 12 cm
pots using planting mix purchased from Michigan Peat Company, Houston, TX, and
placed in different growth chambers held at either 18, 22, 26, or 30 °C and 12 h days and
~600 mo] 5'1 m'2 PAR (7 replicates of each genotype in for each). After a seven week

induction period, the number of flowers and runners were recorded weekly for a ten-week

26

period. At the end of this time, the plants were partitioned into root, crown, leaf and
runners. The dry weights of the plant parts were determined afier holding them at 72 °C
for three days.

The same experiment was repeated in the summer of 2002 (from 4/12/02 to
8/17/02); however, some of the genotypes were dropped and replaced by others. The
genotypes included in the second trial were: F. xananassa 'Aromas', 'Fort Laramie',
'Quinalt', 'Tribute', and elite 'F. virginiana genotypes Frederick 9, LH 50-4, RH 30.
Fragaria virginiana clones Brighton-3, LH 30-4, LH 39-15, LH 40-4, RH 23, RH 43, RH
45 were dropped as they showed little evidence of being heat tolerant in the first trial.
Also, the second trial had 6 replicates for each treatment for each genotype. The number
of flowers, and runners were recorded weekly for 4 weeks after a 7-week induction
period. At the end of the experiment, the total dry weight of each plant was determined
by drying them at 72 °C for three days.

The mean values for each genotype in the two trials are presented separately, as
different genotypes were included in each. However, the common genotypes in each trial
were considered as blocks to calculate Analysis of Variance tables. ANOVAs, means,
and SEs were calculated using the SAS program (SAS Institute, Cary, N.C.). The
regression lines were fitted on average values for each genotype plotted on day length
treatments to demonstrate the trend of the genotype’s performance over day length

treatments.

Results and Discussion

27

There was a significant difference in the two trials for plant weight; however,
number of crowns, inflorescence, flowers and runners were similar in both trials (data not
shown). Plants were 37% (means = 8.5 vs. 6.2 g) heavier in the second trial.

Temperature had a significant effect on flower number (Table 5), but not crown and
inflorescence numbers (data not shown). The overall mean flower number increased
progressively as temperature decreased. The mean values were 1.6, 3.1, 3.9, and 6.8 at
30, 26, 22, and 18 °C (Table 6). F ragaria xananassa cultivars had higher mean flower
numbers than F. virginiana genotypes (5.7 vs. 2.8 overall and 8.4 vs. 5.3, 5.5 vs. 2.5, 4.2
vs. 2.1, and 3.3 vs. 0.4 for 18, 22, 26, and 30 °C, respectively) (Table 6).

Both the main effect of genotype and the genotype x temperature interaction were
highly significant (Table 5). The overall flower number ranged from 0.7 (Frederick 9) to
10.6 (RH43) in F virginiana genotypes and 2.9 ('Fort Laramie') to 9.1 ('Tribute') in the F
xananassa cultivars (Table 6). All of the F. virginiana genotypes, except RH 43, had
their lowest flower numbers at the high temperatures (26 and 30 °C) and showed a non-
significant, negative relationship; the rate of decline was only significant for RH 43
(Table 6 and 7). 'Aromas', and 'Tribute' also had their lowest flower number at 30 °C (1.7
and 2.6, respectively) (Table 6 and Figure 6) and showed a non-significant negative trend
(Table 7). Floral production in LB 'Fort Laramie' however, was higher at 30 °C than 18
°C and it displayed a non-significant positive relationship (Tables 6 and 7).

Temperature did not have a significant effect on runner number, although the
main effect of genotypes and the genotype x temperature interaction were highly
significant (Table 5). LH 40-4, RH 23, and RH 45 did not runner in any of the

temperature treatments, while LH 50-4 produced almost equal runner numbers at all

28

temperatures (overall mean = 2.5) (Table 6). All of the F xananassa cultivars had very
low runner numbers regardless of temperature (Table 6 and Figure 7).

For dry weights, the main effect genotype and the genotype x temperature
interaction were highly significant, while the main effect of temperature was not
significant (Table 5). F ragaria xananassa cultivars had higher dry weights (overall
mean = 9.4) than the F. virginiana genotypes (overall mean = 4.5) (Table 6). Overall, the
mean dry weights of both species were significantly reduced by high temperatures (12.4,
9.9, 9.0, and 6.7 for F. xananassa and 7.5, 3.7, 3.2, and 3.2 for F. virginiana for 18, 22,
26, and 30 °C, respectively) (Table 6 and Figure 8). The smaller plants of F virginiana
had flat, non-significant regression lines, while most of the other genotypes showed non-
significant negative trends as temperature was increased. Only 'Aromas' and 'Tribute' had
negative trends that were significant (Table 7).

Unfortunately, all of the DN genotypes screened were negatively affected by high
temperature. The trends were often non-significant but in all cases flower numbers were
substantially higher at 18 °C than 26 °C and 20 °C. Several other studies have shown
growth rates, flowering and C02 assimilation rates to be reduced in strawberry by high
temperatures and this relationship is probably reflected here in biomass (Hellmann and
Travis, 1998; Serge et al., 2002).

Because the genotypes in the experiment came from a wide range of
environments, there is a diminishing hope of finding a DN genotype that flowers well
under high temperatures. This suggests that in hot northern climates, it may be a better
breeding strategy to concentrate on LD double cropping parents rather than DN multiple

cropping ones. LD genotypes flower in the relatively cool days of spring and late

29

summer, and thus avoid the hot, mid-summer temperatures. In fact, the old everbearing
cultivars 'Fort Laramie' is LD and appears to have some resistance to heat. The fi'uits of
'Fort Laramie' do not meet current commercial standards, but these cultivars could prove

to be a useful parent.

30

Table 5. Analysis of variance for flower and runner numbers, and dry weights of
strawberry genotypes grown in growth chambers at 12 h days and 18, 22, 26, and

 

 

 

30 °C.
Source df Flower no. Runner no. Dry weight (g)
Block 1 55.1 1.0 79.1“
Temperature (T) 3 158.1 ** 1.2 9.5
Whole-plot error 7 13.7 0.7 28.7"
Genotype (G) 14 236.9" 15.1" 220.5M
G*T 40 56.9" 1.3" 20.0“
Error 340 26.1 0.7 8.0

 

*, " Significant at 0.05 and 0.01, respectively.

31

 

 

 

 

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37

CHAPTER 2
GENETICS OF DAY-NEUTRALITY IN F RA GARIA xANANASSA AND

F RA GARM VIRGINIANA

Introduction

Breeders outside of the Mediterranean climates are very interested in identifying
new, much stronger sources of day-neutrality that are also highly resistant to summer
heat. To this end, native strawberry clones have been collected from Alaska, Alberta,
Minnesota New York, the northern Rocky Mountains, Ontario, Pennsylvania, western
North Carolina (Luby et al., 1992; Hancock et al., 1993; Hokanson et al., 1993; Sakin et
al., 1997). Over 2,500 native strawberries have now been evaluated for their flowering
types and horticultural attributes, and out of this group, several elite day-neutral clones
have been selected (Hancock et al., 2001 a and b). These clones originated fi'om a wide
geographical range, including climates with high summer temperature (Hancock et al.,
2001 a and b; www.berrvgenetics.org).

Several different models have been proposed to explain the genetic control of
photoperiod control in strawberries. These include: 1) regulation by a single-gene, 2) two
complementary genes without modifiers, and 3) two complementary genes with
modifiers (Clark, 1927; Powers 1954; Ahmadi et al., 1990). In those models, the genes
regulating day-neutrality have been proposed to be dominant, recessive, or both with no
maternal effects (Macoun, 1924; Clark 1937). Unfortunately, these studies were

conducted using several different genetic sources of day-neutrality.

38

Most researchers consider a strawberry plant to be DN if it can form flower buds
under both long and short day conditions. The most precise method of evaluating day-
neutrality is to monitor the plants during the whole growing season in the field; however,
several less time consuming methods have been used in greenhouses (Ahmadi et al.,
1990). For example, a seedling is considered to be DN if it flowers 3 months after
germination or if there are flowers on new runner plants. It is possible that some of the
discrepancies in the literature on the genetics of day-neutrality have been caused by the
use of different methods of determining day-neutrality. It is also not known how well
field and greenhouse are correlated. Since the use of greenhouse and seedling data is the
fastest way to identify DNs for cultivar development, knowledge about how well the
various methods correlate is critical.

The genetics of day-neutrality has been studied extensively without any consistent
results. It was not until the 1970s that the term DN was employed instead of everbearer,
but any genotype will be considered as DN; if they were shown to flowered under both
short and long days. In the first study done Richardson (1917), European F. xananassa
everbearing mutants ('St. Antonie Padoue' and 'Laxton’s Perpetual') was used to
investigate the genetics of DN. When Richardson made DN x DN and DN x SD crosses
he did not get the expected 3:1 and 1:1 ratios, respectively, if day-neutral was a dominant
trait. His conclusion was that this trait is controlled by a partially dominant gene with
complex interactions.

Using North American F. xananassa mutants that were probably derived from
'Pan American’, Macoun (1924) and Darrow (1937) found DN x DN crosses fit a 9:7 ratio

and SD x DN crosses fit a 3:1 ratio. This is consistent with a dominant, complimentary

39

gene model. Clark (1937) also found similar mean values in DN x DN and DN x SD, but
he uncovered sufficient differences among cultivars in the proportion of DN progeny to
lead him to implicate significant factor interactions. Clark (1937) used North American
F. Xananassa mutants like Macoun (1924) and Darrow (1937), but his source of DN was
probably different as he obtained his day-neutrals from short day 'Mastodon' x short day
'Howard 17' crosses.

Powers (1954) was the first investigator to analyze segregation patterns in DN
material derived from F. virginiana ssp. glauca, as well as North American F. Xananassa
mutants. He concluded that this trait is control by several (at least six) dominant and
recessive genes; and suggested that the dominant genes are not equal in power in
conditioning the expression of day-neutrality. He indicated that there were three
independent loci and ranked their dominant allele based upon their power in conditioning
day-neutral progeny (A’ >A > B and C).

Using a mixture of North American F. Xananassa clones where the DN gene
came from both North American F. Xananassa mutants and F. virginiana ssp. glauca,
Orecky and Slate (1967) also provided progeny data that implicated a complimentary
genes model. However, they detected significant parent specific deviations from the
model and suggested octosomal segregation to explain this variability.

Bringhurst and Voth (1978) examined segregation of DN in populations derived
from F. virginiana ssp. glauca. They found that this trait appeared in about 40% of the
offspring in DN x SD crosses and concluded that DN is controlled by a single dominant
gene. Ahmadi et al. (1990) later confirmed this in crosses of F. Xananassa carrying the

Wasatch source with short day types of F. Xananassa and F. chiloensis. They also found

40

three dominant genes that control the expression of the day-neutrality trait in native
California F. vesca. This is in contrast with work by Brown and Wareing (1965) where
they found day-neutrality in 'Baron Solemacher' and 'Bush white' of European F. vesca to
be controlled by single, recessive gene.

Most recently, Hancock et al. (2002) investigated the inheritance of DN by
crossing SD and DN representatives of F. virginiana to SD and DN F. Xananassa
cultivars. The resulting progeny were evaluated in three different locations (Michigan,
Minnesota, Ontario). When DN F. virginiana genotypes were hybridized with SD F.
Xananassa, they produced ratios which all deviated significantly from a 1:1 ratio
indicating photoperiod sensitivity is not controlled by a single dominant gene. The
percentage of DN progeny produced by each of the day-neutral F. virginiana parents
varied widely in a quantitative fashion and, in fact, a few DN progeny (up to 11.5%) were
covered in crosses of SD F. virginiana x SD F. Xananassa and SD F. Xananassa x SD F.
virginiana. In addition, they detected significant differences in expression of DN among
crosses across locations, with Ontario producing the highest percentage DN progeny.

If all the studies of the genetics of DN are considered together, we must conclude
that the genetics of day-neutrality in strawberries is still unknown. No consistent patterns
of inheritance have emerged. One of the most important reasons for the discrepancy in
the literature is the employment of different sources of DN. F ragaria virginiana ssp.
glauca was used as DN source in different backgrounds (Powers 1954; Orecky and Slate
1967; Bringhurst and Voth 1978), and the F. xananassa sources of DN also varied
greatly. Richardson (1917) used European source came from 'F. de Gaillon', while

Macoun (1924) and Darrow (1966) used North American source, 'Pan American', which

41

was a chance seedling of 'Bismark’. Clark (1937) used a DN source derived fi'om a cross
of SD 'Mastodon' x SD 'Howard 17'. In fact, there have been more than six independent
and original sources of DN identified in F. xananassa and transferred to SD types
(Darrow, 1966).

It is also possible that differences in evaluation methods and timing have
contributed to the variation in segregation ratios. Numerous methods of screening DN
have been employed including 1) flowering on mother and runner plants during one
summer and fall, 2) flowering patterns across two seasons, 3) how fast seedlings flower,
and 4) crossing individuals to SD F. chiloensis and analyzing the percentage of day-
neutral progeny produced (Nicoll and Galletta, 1987; Ahmadi et a1. (1990). It is not
known how tightly these evaluation methods are associated. Timing of evaluation is
critical as well. Ahmadi et al. (1990) noted that SD genotypes with little chilling
requirement might initiate flower buds in August and flower in November. If the
progeny were scored during that time, the genotype could have been misjudged as DN.
Also, SD genotypes flower semi-continuously in the second year in mild climates. The
various investigators who have studied the genetics of day-neutrality rarely used the same
dates. For example, Richardson (1917) scored his genotypes from May to October, while
Powers (1954) evaluated them fiom July to September.

The use of the term “everbearing” has added complexity to the literature
regarding the genetics of day-neutrality. Everbearers have been variously described as a
plant fi'uiting more than one time in a year and synonymies as “perpetual”, “four-season”,
“rebloomer”, “remontant”, “double-cropping”, and finally “day-neutral” (Galletta and

Bringhurst, 1990). Nicoll and Galletta (1987) stated that the term “day-neutral” and

42

“everbearer” can be used interchangeably, if DN is used as to a physiological term to
denote a relative insensitivity to day length in flower bud initiation and everbearer is used
as an agricultural term to indicate a pragmatic expectation of summer strawberry
production. However, the distinction should be made clear between DN and everbearer
in genetic studies since both the DN and LD physiological classes have been referred to
as everbearer in the literature.

In this chapter, experiments were designed to elucidate the genetics of day-
neutrality in native clones of F ragaria, to more efficiently utilize them in breeding new
day-neutral F ragaria xananassa cultivars. The specific objectives were to determine: 1)
the relationship between several different evaluation methods for DN, 2) if greenhouse
screens can be used to predict field flowering performance, 3) if day-neutrality is a
quantitative or qualitative trait, 4) if there is more than one source of genes for day-
neutrality in native North American populations of F. virginiana, and 5) if genetic

background has an influence on the expression of day-neutrality.

EXPERIMENT 1

Material and Methods

This experiment was designed to evaluate different methods of identifying DN

progeny. Segregating populations were constructed in a partial-diallel fashion (Table 8).

Information on F. virginiana clones can be found at Sakin et al. (1997), Hancock et a1.

43

(2001 a and b) and www.berrygenetics.com. DHL 1336 ('Tribute' x Montreal River 10)
is a selection from the MSU Strawberry Genetics Program.

All parental genotypes were potted onto 18 x 16 x 13 cm pots in the summer of
1999) and placed in a greenhouse at MSU with natural day lights and a mean temperature
of ~ 21 °C. Crosses were made by transferring pollen with a camel hairth after
removal of stamens using sharp tweezers to prevent self-pollination. Fresh pollen was
generally collected from open flowers; however in some occasions, pollen was stored in
petri dishes at -16 °C for future use. The fruits were harvested when fully ripe and seeds
were extracted by smashing the fruits on paper towels. To promote germination, seeds
were then placed on soil in pots and held in a grth chamber at 4 °C with continuous
inflorescent light and moisture. In February 2000, when the seeds started to germinate,
they were placed in a growth room with ~ 18 °C with continuous light.

Each seedling was evaluated using five methods to score day-neutrality: 1) They
were planted in March 2000 into 14 x 12 x 12 cm pots and placed in a greenhouse at
MSU under long day conditions (13 h days created with ~ 800 mo] 8'1 m'2 of
supplementary light) and 18 - 22 °C. Any genotype that flowered within 100 days from
germination was considered DN (DN1-100 days to flowering). 2) The seedlings were
transplanted in the field at the Southwestern Michigan Research and Extension Center

(SWMREC) on 07/25/2000 at 60 x 120 cm spacing and any that flowered in that same

44

Table 8. The genotypes crossed in a partial-diallel fashion to study the interaction of
different evaluation method of day-neutrality.

 

 

 

 

 

F ragaria xananassa F ragaria virginiana
Day-neutral or
everbearer Short day Day-neutral Short day
'Aromas' DHL 1336 Frederick 9 Eagle 14
'Fort Laramie' 'Camarosa' LH 39-15 High Falls 22
'Ogallala' 'Honeoye' RH 30 Montreal River 10
'Tribute' 'Glooscap' RH 18

 

4S

summer before 9/9/2000 were considered DN (DN2-1St year field flowering). 3) Rooted
one-year-old runner plants were collected from each of these mother plants and placed in
an unheated greenhouse in September 2000, and allowed to flower in the spring of 2001
without supplementary light. Those plants that flowered again before 9/01/01 were
considered as DN (DN3-2“d year greenhouse flowering). 4) All of the original field-
grown plants were monitored another year for flowering in the spring and summer of
2001. Those that flowered in the spring and again before 9/9/2000 were considered as
DN (DN4-2"d year field flowering). 5) During this same period in the field, genotypes
that produced flowers on their newly formed runners were considered DN (DNS-runner
flowering). In all experiments, each family was maintained in a single plot without
replication.

Greenhouse surveys can only be used to identify elite DN progeny if the
photoperiod response of progeny is similar in the greenhouse and field, or if there are
fewer DN progeny in the greenhouse, those that do produce multiple greenhouse crops
are at least the strongest DN genotypes in the field. To investigate this possibility, each
field grown genotype was given a flowering strength rating (F SR) of 0 - 10 in the
summer of 2001 , alter its photoperiod sensitivity had been rated in the 2000 greenhouse
screens. These plants with no flowers were rated 0, and those with the most were given a
10.

A number of horticulturally important traits were also evaluated for each of the
hybrids in the summer of 2001. Crown and runner numbers were counted on 5/6/01, and
inflorescence and flowers per inflorescence were recorded on 5/9/01. Four fruits from

each plant were harvested on 6/13/01 and weighed to calculate average fruit weight.

46

The percentage of DN progeny was calculated for each family and the grand mean
for each family using each evaluation method was determined. Correlations among these
scores were calculated using the SAS program (SAS Institute, Cary, N.C.). The mean

and SEs of the horticulturally important traits were also determined for each family.

Result and Discussion

Dav-neutrality evaluation methods: Different mean percentages of DNs were
observed across the evaluation methods. DNS (runner flowering) had the lowest overall
mean (18%), while DNl (100 day flowers) had the highest (55%) (Table 9). The
greenhouse evaluations produced higher means than the field evaluations (55 and 49%
vs. 41 and 40%). Large amounts of variation were observed across families in the mean
% DN progeny for each method. In fact, in three of the evaluation methods, the range in
family values was 0 - 100% (DNl, 100 days flowers, DN3 - 2‘“l year field flowering,
DN4 - 2'“1 year greenhouse flowering). DN x DN crosses generated higher numbers of
DN progeny than DN x SD crosses for all methods. For example, when DN 'Aromas'
was selfed it produced 60, 67, 67, 100, and 33% for DNl, DN2, DN3, DN4, and DNS,
respectively, while SD ‘Glooscap' x SD RH 18 produced 47, 38, 27, 6, and 0 % (Table 9).

Some of the crosses with high numbers of DN progeny in the field and greenhouse
screens did not have any flowers on their runners; for example, 'Aromas' x 'Fort Laramie',
and 'Aromas' x DHL 1336 (Table 9). The highest family values for flowers on their
newly-formed runners (DNS-runner flowering) were 64, 39, and 38% for 'Tribute' x

Frederick 9 (64%), 'Tribute' x RH 18 (39%), and 'Tribute' x Eagle-14 (38%) (Table 9).

47

'Tribute', which usually forms flowers in its runners, is the common parent in these
crosses (Draper et al., 1981; Maas and Cathey, 1987).

The average flowering strength rating (F SR) across all families was 1.2 (Table 9).
Eagle-14 x Eagle-14 had the lowest average F SR (mean = 0.0, N = 28), while 'Aromas' x
'Ogallala' (mean = 3.8, N = 9), 'Tribute' x RH 18 (mean = 3.4, N = 38), and 'Tribute' x
Eagle-14 (mean = 3.4, N = 39) had the highest FSR rating.

Hundred-day flowering (DNl) was significantly correlated with DN2 (1St year field
flowering) (R = 46%, P = 0.003), but not with any of the other methods (Table 10).
However, all the other evaluation methods were significantly correlated (Table 10 and
Figure 9 and 10). Likewise, the FSR were significantly correlated with all the DN
evaluation methods except DNl (flowering within 100 days) (Table 10). The highest
correlation was observed between DN4, 2ud year field flowering, and FSR. This is not
surprising, as these data were collected at the same time in the field and a high value for
strength of flowering also indicates the plants are strong day-neutrals (Anonymous,
1988). Also, the year-to-year correlation in the field (DN4 vs. DN2) was high (71%, P =
0.000), and the greenhouses vs. field evaluations in 2001 were highly significant (73%, P
= 0.000 (Table 10 and Figure 9).

Horticulturaflraits: The families had quite variable averages for all of the
horticulturally important traits (Table 11). Mean crown number ranged from 2.1
('Aromas' x 'Ogallala' and 'Aromas' x 'Tribute') to 6.4 ('Fort Laramie' x 'Frederick 9')
(Table 11). 'Aromas' x 'LH 39' did not have any runners, while Eagle-14 x Eagle-14 had
very high runner numbers (11.5) (Table 11). 'Aromas' x 'Tribute' produced the lowest

inflorescence and crown numbers (mean = 4.4), while DHL 1336 x 'Glooscap' had the

48

highest numbers (mean = 12.5) (Table 11). The range in flowers per inflorescence was
from 3.1 ('Aromas' x 'Ogallala') to 6.7 ('Ogallala' x 'Camarosa') (Table 11). The average
fruit weight in Eagle-14 x Eagle-14 was extremely low (mean = 1.4), while 'Camarosa' x
’Glooscap' had the largest fruits (mean = 13.8). Among these traits, only runner number
was negatively associated with % DN in the families (Table 12).

Conclusions: These results indicate that scoring DN progeny within 100 d from
germination can not be used to predict field performance. Apparently, the speed with
which a seedling begins flowering is not tightly associated with photoperiod sensitivity.
However, greenhouses can be used to predict field performance, if the flowering behavior
of individuals is followed through a whole season. The DN percentage observed in our
second year greenhouse screens were highly correlated with the subsequent field
evaluations, and the families with the highest flowering strength in the field also had the
highest percentage of DNs in both the greenhouse and field screens.

The final decision on whether greenhouses will be utilized in a breeding program
will still have to be based on the objective of the breeding program. While the
correlation between the field and greenhouse determinations of % DN in families was
high, some families which had no DN individuals in the greenhouse, did show some in
the field. For example, the second year greenhouse evaluations of DHL 1336 x
'Honeoye' crosses generated no DNs, while 17% were DN were recovered in the field. If
one is interested in finding DNs in the broadest range of families, the populations need to
be evaluated in the field, probably for multiple years. Additionally, the relationship
between DN and number of linking cycles was not evaluated. If numbers of cycles is

important, populations will need to be screened directly for that characteristic.

49

Small fruit size is a common problem in DN breeding and it has been suggested that
fruit size and day-neutrality are negatively correlated (Dale et al., 2002). We did not find
this to be the case in this study (R2= 0.18, df= 39, P = 0.275) (Table 12). In fact, the
only significant correlation observed between day-neutrality and horticulturally important
traits was a negative correlation with runner number (- 49%, P = 0.001) that has been
previously demonstrated (Hancock et al., 2002). This suggests that with the exception of
runner numbers, there are a few negative compensation that will impede DN breeding
using the populations in this study. Even though runner number was negatively
correlated with %DN, some families produced runners than the current DN cultivars

indicating that improved runnering types can be recovered.

50

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52

Table 10. Correlation coefficients, significance, and number of individuals in strawberry
families grown in a greenhouse at Michigan State University, East Lansing, Mich.,
and in the field at the Southwest Michigan Research and Extension Center Benton
Harbor, Mich. in 2000 and 2001.

 

 

 

 

 

 

1)sz DN33 DN44 DN55 FSR6

131511I 0.46 0.26 0.27 0.16 0.06
0.0037 0.125 0.095 0.320 0.726

£39) (36) (39) (39) 139)

DN2 0.71 0.71 0.33 0.54
0.000 0.000 0.042 0.000

(36) (39) (39) (39)

DN3 0.73 0.45 0.74
0.000 0.005 0.000

(36) (36) (36)

DN4 0.70 0.85
0.000 0.000

(39) (39)

DNS 0.58
0.000

(39)

 

IF lowering within 100 days from germination in a greenhouse in 2000.

2F lowering before 9/9 in field in 2000.

3Flowering under both short and long days in a greenhouse in 2001.

‘Flowering under both short and long days in field in 2001.

sFlowering on their newly-formed runner in field in 2001.

6Flowering strength ratio of 0-10 (10 having the most flowers during the second cycle of flowering) in field

in2001.

7Sigfificant P values, at 0.05, are bolded.

53

Figure 9. Association of several day-neutrality scoring methods in strawberry families
grown in a greenhouse at Michigan State University, East Lansing, Mich., and in
the field at the Southwest Michigan Research and Extension Center Benton Harbor,
Mich. (DNl-Flowering within 100 days from germination in a greenhouse in 2000;
DN2-Flowering before 9/9 in field in 2000; DN3-F lowering under both short and
long days in a greenhouse in 2001; DN4-Flowering under both short and long days

 

 

 

    

in field in 2001).
y = 0.85x + 5.1 R1 . 0.51 y = 0.23x + 33.6 R2 . 0.05
100 .
80 g
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Greenhouse 2001 (DN3)

54

Figure 10. Association of several day-neutrality scoring methods in strawberry families
grown in a greenhouse at Michigan State University, East Lansing, Mich., and in
the field at the Southwest Michigan Research and Extension Center Benton Harbor,
Mich. (DN4-Flowering under both short and long days in field in 2001; DNS-
Flowering on their newly-formed runner in field in 2001; Flowering strength ratio
of 0-10 (10 having the most flowers during the second cycle of flowering) in field
in 2001).

 

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55

 

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57

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EXPERIMENT 2

Material and Methods

To investigate the genetics of day-neutrality, two groups of segregating populations
were constructed using a diallel design: 1) DN x SD crosses, and 2) DN x DN crosses
(Table 13). An elite DN F. virginiana genotype, RH 23 (Hancock et al., 2002) was also
crossed to some genotypes. Crosses were made as previously described in Experiment 1.
The fruit were harvested and seed were extracted and placed on soil in pots and held in a
grth chamber at 4 °C with continuous inflorescent light. When the seeds germinated
in summer 2001, the seedlings were placed in a growth room ~18 °C with continuous
inflorescent light (~600 mol 3'1 m'z). When they had reached the 4 - 6 leaf stage, they
were potted into 14 x 12 x 12 cm pots and placed in a greenhouse at ~18 °C and 13 h
long day conditions maintained by high intensity lamps (~800 umol s'l m’z). Plants
began to flower on 01/28/02 and by mid-April all flowering had ceased. In May 2002,
segregating populations were planted at the MSU Horticulture Farm in a completely
randomized design. Each family was divided into four replications and the row spacing
was 180 cm x 60 cm. Flowering was then monitored on a weekly basis from July 26 to
August 24 in the field. Similar to DN4 (2“‘1 year field flowering) of Experiment 1,
genotypes were considered as DN if they flowered under the short days of spring before

5/30/02 (<14 h) and the long days of summer after 7/24/02 (>15 h).

59

Table 13. The family numbers of genotypes crossed in a partial-diallel fashion to study
the genetic of day-neutrality in strawberries.

 

 

 

 

 

 

 

Day-neutral
F. xananassa F. virginiana
Genotype 'Aromas' 'Fort Laramie' 'Tribute' Frederick 9 LH 50—4 RH 23 RH 30
Short day
F. xananassa
'Allstar' 53 86 89 52
'Chandler' 53 71 72 36 89
'Honeoye' 71 54 72
F. virginiana
Eagle-14 71 54 54
FRA 0368 54 29 72 53
NR 10 72
Day-neutral
F. xananassa
'Aromas' 18 70
'Fort Laramie' 30 54
'Tribute' 52 36 72 71
F. virginiana
Frederick 9 53 72
LH 50-4 36 51
RH 23
RH 30

 

60

Chi-square tests were constructed for each family using either a 1:1 (DN x SD crosses) or
3:1 ratio (DN x DN crosses), assuming DN was regulated by a single dominant gene
(Ahmadi et al., 1990). The tests were also made using pooled data (all DN x all DN, all
DN F. virginiana x all DN F. virginiana, and all DN F. virginiana x all DN F
xananassa).

F ragaria xananassa x F. virginiana in both DN x SD and DN x DN groups were
combined to construct Analysis of Variance tables to test the significance of general and
specific combining abilities using the SAS GLM procedure (SAS, 1990). To increase
- normality, the percentage of day-neutral progeny in each family was m
transformed, although the means are presented as untransformed data. Both F.

xananassa and F. virginiana genotypes were considered as fixed in the analysis.

Results and Discussion

Chi-square analyses: Overall, 58% of the progeny were DN in the combined DN x
SD crosses (Table 14). This percentage is significantly higher than the 50% that is
expected if DN is regulated by a single dominant gene. The F. xananassa x F
xananassa (59%) and F. xananassa x F. virginiana (62%) crosses also averaged
significantly more DN progeny than expected under the single—gene model. The average
percentage of DN progeny in the F. virginiana x F virginiana crosses (48%) did not vary
significantly fi'om a 1:1 (Table 14).

Most of the individual families did not fit the single gene model for the inheritance

of DN. A continuous variation in % DN progeny in the F xananassa x F. virginiana

61

families was observed, ranging from 30% ('Allstar' x LH 50-4) to 87% ('Tribute' x Eagle-
14) (Table 14 and Figure 11). All but one of the F. virginiana x F. virginiana families
(F RA 0368 x LH 50-4, 36%) differed significantly fi'om the expected 1:1 ratio (Table
14). The F. xananassa x F xananassa families generally fit a 1:1 model except 'Allstar'
x 'Fort Laramie' and 'Tribute' x 'Chandler'.

Overall, 70% of the progeny were DN in the combined DN x DN crosses; which is
significantly lower than the expected 75% (3:1 ratio) under the single, dominant gene
model (Table 15). While the mean of the F. xananassa x F xananassa crosses did not
differ significantly fiom the expected values (76% vs. 75%), the average percentage of
DN in the F. xananassa x F virginiana crosses was significantly higher (83%) and the F.
virginiana x F. virginiana crosses was significantly lower (48%) than expected (Table
15). In particular, the selfing of 'Tribute' produced extremely high numbers of DN
progeny (88%) and the highest number of DN for any family was observed in 'Tribute' x
RH 23 (93%). All crosses in the F. virginiana x F. virginiana group had significantly
lower numbers of DN progeny than expected (<63%) (Table 15). Similar to the DN x SD
families, the DN x DN families displayed continuous variation in their percentage of DN
progeny (Figure 12).

General and Smcific Combining Abilities: The analysis of variance indicated that
general combining ability (GCA) for DN was significant in both E xananassa and F.
virginiana (Table 16). Likewise, specific combining ability (SCA) was significant (P =
0.000) in both species. The largest proportion of the variance component was in SCA
(34%), while GCAF, xamsa and GCAF, ngm tabulated 22 and 20% of the variation,

respectively (Table 16). A continuous pattern of variation in parental means was

62

observed for the percentage of day-neutral progeny produced, ranging fi'om 55.1%
('Honeoye') to 85.0% ('Tribute') among F. xananassa cultivars and from 49.5% (LH 50-
4) to 76.0% (Eagle-14) in F. virginiana genotypes (Figure 13).

Conclusions: Such a wide range in the percentage of DN progeny found in the
various families suggests that day-neutrality in octoploid strawberries is not regulated by
a single, dominant gene. Several different kinds of observations support this conclusion.
1) Less than half the families produced 1:1 or 3:1 ratios of day-neutral plants. 2) DN F.
virginiana genotypes produced significantly different percentages of DN progeny than
DN F. xananassa cultivars. 3) Two different DN parents crossed to the same short day
genotype produced different percentages of DN progeny. For example, when ’Tribute'
and 'Aromas' were crosses to the same SD genotype, 'Tribute' consistently generated
more DN progeny than 'Aromas'. 4) Some of the day-neutrality sources were more
powerful than others in producing of day-neutral progeny (e. g., 'Tribute' > 'Aromas' and
RH 23 > Frederick 9). Both general and specific combining abilities for DN were
significant. 5) None of the DN parents produced 100% DN progeny, which would be
expected if there were homozygous dominant DN individual. Repeated selfing of DN
parents has increased the percentage of DN progeny, but no true DN breeding genotypes
have been produced (Shaw, personal communication). 6) In a previous study, SD x SD
crosses generated some (up to 8.2%) DN progeny (Hancock et al., 2002). Such SD
genotypes were not expected from SD x SD crosses under the single, dominant gene
model.

In the literature reporting the genetics of the everbearing trait in strawberries, only

the latest paper fiom the University of California-Davis suggests that multiple cropping is

63

regulated by a single dominant allele at a single nuclear gene locus (Ahmadi etal., 1990).
The other studies do not always distinguish between DN and LD sources of the
everbearing trait, but they uncover a wide array of segregation patterns that did not fit a
single, dominant gene model (Hancock, 1999). The segregating populations that
Bringhurst’s group used to test their genetic hypothesis must have difi‘ered fi'om all the
others in that it contained a unique source of DN from them or it was fixed for the other
quantitative trait loci (QTL) associated with photoperiod sensitivity. All their DN parents
must have been heterozygous.

In this study with a broader range of germplasm, the inheritance of DN appeared
quantitative with quite variable numbers of DN being produced from the various parental
combinations. Similar results were observed in another study, when crosses between DN
F. xananassa and DN F. virginiana fit a 3:1 ratio in only 3 out of 10 combinations
(Hancock et al., 2002). When Barritt et al. (1982) tested the genetics of day-neutrality
using the eastern genotypes, they also obtained complex ratios, as did Powers (1954)
when he examined segregation ratios in a population that contained DN genes from both
F. xananassa 'Pan American' and F. virginiana ssp. glauca. Similar to results of present
study, Powers found continuous variation in % DN progeny and suggested two or more
dominant genes and at least four recessive genes regulating expression of day-neutrality.
Powers also suggested that the effects of dominant alleles are not equal and the effects of
the genes are cumulative.

It is possible that some of the variation we recovered among families for percent
DN progeny was created by the genomic structure of strawberry. The single, dominant

gene model was tested assuming complete diploidization of strawberry genome;

64

however, the pairing relationship octoploid strawberries have not been completely
resolved. Strawberry is considered to be an autopolyploid (Hancock, 1999) so variation
in meiotic configurations of chromosomes might be expected, resulting in occasional
aberrant segregation ratios. Arulsekar and Bringhurst (1981) used isozyme data to
document disomic inheritance in strawberries, but they only evaluated a few loci.

In conclusion, the results indicate that the genetics of day-neutrality is quantitative
in strawberry. The numerous investigators who obtained different segregation ratios
were probably studying different QTLs controlling day-neutrality and in many instances
were using different evaluation methods. It is recommended that quantitative approaches
be utilized in breeding day-neutral strawberries. To produce families with the highest
proportion of DN progeny, parents should be selected which carry the highest number of

QTL for DN.

65

Table 14. Proportion of short day (SD) and day-neutral (DN) progeny generated in SD x
DN crosses. Chi-square tests were made assuming DN was regulated by a single
dominant gene (1 :1). The families were grown in a completely randomized design
at the Michigan State University Horticulture Farm, East Lansing, Mich. in 2002.

 

 

 

 

 

 

 

 

1?ng SD DN Total %DN x2 P
F. xananassa x E xananassa
'Tribute' x 'Allstar' 46 40 86 0.47 0.4 0.518
'Aromas' x 'Chandler' 24 29 53 0.55 0.5 0.492
'Tribute' x 'Honeoye' 30 41 71 0.58 1 .7 0.192
'Chandler' x 'Fort Laramie' 30 41 71 0.58 1.7 0.192
'Allstar' x 'Fort Laramie' 19 34 53 0.64 4.2 0.0391
'Tribute' x 'Chandler' 19 53 72 0.74 16.1 0.000
Total 168 238 406 0.59 12.1 0.001
E Xananassa x F. virginiana .
'Allstar' x LH 50-4 62 27 89 0.30 13.8 0.000
'Tribute' x FRA 0368 17 12 29 0.41 0.9 0.353
'Honeoye' x LH 50-4 38 34 72 0.47 0.2 0.637
'Honeoye' x Frederick 9 26 28 54 0.52 0.1 0.785
'Chandler' x RH 30 36 53 89 0.60 3.2 0.072
'Allstar' x RH 23 21 31 52 0.60 1.9 0.166
'Chandler' x RH 23 14 22 36 0.61 1.8 0.182
'Honeoye' x RH 30 25 47 72 0.65 6.7 0.010
'Fort Laramie' x Eagle-14 23 48 71 0.68 8.8 0.003
'Chandler' x LH 50-4 20 51 71 0.72 13.5 0.000
'Aromas' x F RA 0368 13 41 54 0.76 14.5 0.000
'Allstar' x Frederick 9 11 60 71 0.85 33.8 0.000
'Tribute' x Eagle-14 7 47 54 0.87 29.6 0.000
Total 313 501 814 0.62 43.4 0.000
E virginiana x F. virginiana
FRA 0368 x LH 50-4 46 26 72 0.36 5.6 0.018
Eagle-14 x Frederick 9 31 23 54 0.43 1.2 0.276
FRA 0368 x RH 23 25 28 53 0.53 0.2 0.680
Montreal River 10 x Frederick 9 29 43 72 0.60 2.7 0.099
Total 131 120 251 0.48 0.5 0.487
Grand total 612 859 1471 0.58 41.5 0.000

 

rP values indicating significant variation from a 1:1 model, at 0.05, are bolded.

66

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Table 14. Proportion of short day (SD) and day-neutral (DN) progeny generated in SD x
DN crosses. Chi-square tests were made assuming DN was regulated by a single
dominant gene (1:1). The families were grown in a completely randomized design
at the Michigan State University Horticulture Farm, East Lansing, Mich. in 2002.

 

 

 

 

 

 

 

 

Family so DN Total %DN 2:2 P
E Xananassa x F. Xananassa ,
'Tribute' x 'Allstar' 46 40 86 0.47 0.4 0.518
'Aromas' x 'Chandler' 24 29 53 0.55 0.5 0.492
'Tribute' x 'Honeoye' 30 41 71 0.58 1.7 0.192
'Chandler' x 'Fort Laramie' 30 41 71 0.58 1.7 0.192
'Allstar' x 'Fort Laramie' 19 34 53 0.64 4.2 0.039‘
'Tribute' x 'Chandler' 19 53 72 0.74 16.1 0.000
Total 168 238 406 0.59 12.1 0.001
F. Xananassa x F. virginiana
'Allstar' x LH 50-4 62 27 89 0.30 13.8 0.000
'Tribute' x FRA 0368 17 12 29 0.41 0.9 0.353
'Honeoye' x LH 50-4 38 34 72 0.47 0.2 0.637
'Honeoye' x Frederick 9 26 28 54 0.52 0.1 0.785
'Chandler' x RH 30 36 53 89 0.60 3.2 0.072
'Allstar' x RH 23 21 31 52 0.60 1.9 0.166
'Chandler' x RH 23 14 22 36 0.61 1.8 0.182
'Honeoye' x RH 30 25 47 72 0.65 6.7 0.010
'Fort Laramie' x Eagle-14 23 48 71 0.68 8.8 0.003
'Chandler' x LH 50-4 20 51 71 0.72 13.5 0.000
'Aromas' x FRA 0368 13 41 54 0.76 14.5 0.000
'Allstar' x Frederick 9 11 60 71 0.85 33.8 0.000
'Tribute' x Eagle-14 7 47 54 0.87 29.6 0.000
Total 313 501 814 0.62 43.4 0.000
F. virginiana x F. virginiana
FRA 0368 x LH 50-4 46 26 72 0.36 5.6 0.018
Eagle-14 x Frederick 9 31 23 54 0.43 1.2 0.276
F RA 0368 x RH 23 25 28 53 0.53 0.2 0.680
Montreal River 10 x Frederick 9 29 43 72 0.60 2.7 0.099
Total 131 120 251 0.48 0.5 0.487
Grand total 612 859 1471 0.58 41.5 0.000

 

1P values indicating significant variation fiom a 1:1 model, at 0.05, are bolded.

68

 

 

 

 
 

 

 

 

 

 

 

 

 

 

 

 

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69

Table 16. General and specific combining ability (GCA and SCA) for percent day-neutral
progeny in F. xananassa x F. virginiana families grown in a completely randomimd
design at the Michigan State University Horticulture Farm, East Lansing, Mich. in

 

 

2002.
Mean Variance component
Source df squares F value P anipercentage
GCA (E xananassa) 5 928.5 5.55 0.0011 107.2 (22%)
GCA (R virginiana) 4 993.7 5.94 0.001 100.3 (20%)
SCA 6 914.4 5.47 0.000 165.1 (34%)
Error 42 167.2 117.7 (24%)

 

TSignificant P values, at 0.05, are bolded.

7O

Figure 13. Percentage of day-neutral progeny in strawberry families grown in a completely
randomized design at the Michigan State University Horticulture Farm, East Lansing,
Mich. in 2002. The white column represents F. xananassa cultivars, while black
columns represent F. virginiana genotypes. The bars represent standard errors.

100 1
l

901

801 .{_ {1
701 F}
601 ‘P 71'
50~+

40..
30—

201
101

% DN progeny

 

 

 

 

 

 

 

 

 

 

 

 

 

 

—1
H
_..

'Honeoye'
'Allstar'
'Chandler'
'Ft. Laramie'
'Aromas'
'Tribute'
LH 50-4
Frederick 9
RH 30

RH 23
Eagle 14

71

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