# Patent

## Title
Squash hybrid SV0116YL and parents thereof

## Abstract

The invention provides seed and plants of squash hybrid SV0116YL and the parent lines thereof. The invention thus relates to the plants, seeds and tissue cultures of squash hybrid SV0116YL and the parent lines thereof, and to methods for producing a squash plant produced by crossing such plants with themselves or with another squash plant, such as a plant of another genotype. The invention further relates to seeds and plants produced by such crossing. The invention further relates to parts of such plants, including the fruit and gametes of such plants.

## Background
CROSS-REFERENCE TO RELATED APPLICATIONS
This application claims the priority of U.S. Provisional Appl. Ser. No. 61/846,546, filed Jul. 15, 2013, the entire disclosure of which is incorporated herein by reference.


FIELD OF THE INVENTION
The present invention relates to the field of plant breeding and, more specifically, to the development of squash hybrid SV0116YL and the inbred squash lines ZGN-EH10004 and LEB-EH-08-874.
BACKGROUND OF THE INVENTION
The goal of vegetable breeding is to combine various desirable traits in a single variety/hybrid. Such desirable traits may include any trait deemed beneficial by a grower and/or consumer, including greater yield, resistance to insects or disease, tolerance to environmental stress, and nutritional value.
Breeding techniques take advantage of a plant's method of pollination. There are two general methods of pollination: a plant self-pollinates if pollen from one flower is transferred to the same or another flower of the same plant or plant variety. A plant cross-pollinates if pollen comes to it from a flower of a different plant variety.
Plants that have been self-pollinated and selected for type over many generations become homozygous at almost all gene loci and produce a uniform population of true breeding progeny, a homozygous plant. A cross between two such homozygous plants of different genotypes produces a uniform population of hybrid plants that are heterozygous for many gene loci. Conversely, a cross of two plants each heterozygous at a number of loci produces a population of hybrid plants that differ genetically and are not uniform. The resulting non-uniformity makes performance unpredictable.
The development of uniform varieties requires the development of homozygous inbred plants, the crossing of these inbred plants, and the evaluation of the crosses. Pedigree breeding and recurrent selection are examples of breeding methods that have been used to develop inbred plants from breeding populations. Those breeding methods combine the genetic backgrounds from two or more plants or various other broad-based sources into breeding pools from which new lines and hybrids derived therefrom are developed by selfing and selection of desired phenotypes. The new lines and hybrids are evaluated to determine which of those have commercial potential.
SUMMARY OF THE INVENTION
In one aspect, the present invention provides a squash plant of the hybrid designated SV0116YL, the squash line ZGN-EH10004 or squash LEB-EH-08-874. Also provided are squash plants having all the physiological and morphological characteristics of such a plant. Parts of these squash plants are also provided, for example, including pollen, an ovule, scion, a rootstock, a fruit, and a cell of the plant.
In another aspect of the invention, a plant of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 comprising an added heritable trait is provided. The heritable trait may comprise a genetic locus that is, for example, a dominant or recessive allele. In one embodiment of the invention, a plant of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 is defined as comprising a single locus conversion. In specific embodiments of the invention, an added genetic locus confers one or more traits such as, for example, herbicide tolerance, insect resistance, disease resistance, and modified carbohydrate metabolism. In further embodiments, the trait may be conferred by a naturally occurring gene introduced into the genome of a line by backcrossing, a natural or induced mutation, or a transgene introduced through genetic transformation techniques into the plant or a progenitor of any previous generation thereof. When introduced through transformation, a genetic locus may comprise one or more genes integrated at a single chromosomal location.
The invention also concerns the seed of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874. The squash seed of the invention may be provided as an essentially homogeneous population of squash seed of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874. Essentially homogeneous populations of seed are generally free from substantial numbers of other seed. Therefore, seed of hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 may be defined as forming at least about 97% of the total seed, including at least about 98%, 99% or more of the seed. The seed population may be separately grown to provide an essentially homogeneous population of squash plants designated SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874.
In yet another aspect of the invention, a tissue culture of regenerable cells of a squash plant of hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 is provided. The tissue culture will preferably be capable of regenerating squash plants capable of expressing all of the physiological and morphological characteristics of the starting plant, and of regenerating plants having substantially the same genotype as the starting plant. Examples of some of the physiological and morphological characteristics of the hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 include those traits set forth in the tables herein. The regenerable cells in such tissue cultures may be derived, for example, from embryos, meristems, cotyledons, pollen, leaves, anthers, roots, root tips, pistils, flowers, seed and stalks. Still further, the present invention provides squash plants regenerated from a tissue culture of the invention, the plants having all the physiological and morphological characteristics of hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874.
In still yet another aspect of the invention, processes are provided for producing squash seeds, plants and fruit, which processes generally comprise crossing a first parent squash plant with a second parent squash plant, wherein at least one of the first or second parent squash plants is a plant of squash line ZGN-EH10004 or squash LEB-EH-08-874. These processes may be further exemplified as processes for preparing hybrid squash seed or plants, wherein a first squash plant is crossed with a second squash plant of a different, distinct genotype to provide a hybrid that has, as one of its parents, a plant of squash line ZGN-EH10004 or squash LEB-EH-08-874. In these processes, crossing will result in the production of seed. The seed production occurs regardless of whether the seed is collected or not.
In one embodiment of the invention, the first step in “crossing” comprises planting seeds of a first and second parent squash plant, often in proximity so that pollination will occur for example, mediated by insect vectors. Alternatively, pollen can be transferred manually. Where the plant is self-pollinated, pollination may occur without the need for direct human intervention other than plant cultivation.
A second step may comprise cultivating or growing the seeds of first and second parent squash plants into plants that bear flowers. A third step may comprise preventing self-pollination of the plants, such as by emasculating the flowers (i.e., killing or removing the pollen).
A fourth step for a hybrid cross may comprise cross-pollination between the first and second parent squash plants. Yet another step comprises harvesting the seeds from at least one of the parent squash plants. The harvested seed can be grown to produce a squash plant or hybrid squash plant.
The present invention also provides the squash seeds and plants produced by a process that comprises crossing a first parent squash plant with a second parent squash plant, wherein at least one of the first or second parent squash plants is a plant of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874. In one embodiment of the invention, squash seed and plants produced by the process are first generation (F1) hybrid squash seed and plants produced by crossing a plant in accordance with the invention with another, distinct plant. The present invention further contemplates plant parts of such an F1 hybrid squash plant, and methods of use thereof. Therefore, certain exemplary embodiments of the invention provide an F1 hybrid squash plant and seed thereof.
In still yet another aspect, the present invention provides a method of producing a plant derived from hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874, the method comprising the steps of: (a) preparing a progeny plant derived from hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874, wherein said preparing comprises crossing a plant of the hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 with a second plant; and (b) crossing the progeny plant with itself or a second plant to produce a seed of a progeny plant of a subsequent generation. In further embodiments, the method may additionally comprise: (c) growing a progeny plant of a subsequent generation from said seed of a progeny plant of a subsequent generation and crossing the progeny plant of a subsequent generation with itself or a second plant; and repeating the steps for an additional 3-10 generations to produce a plant derived from hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874. The plant derived from hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 may be an inbred line, and the aforementioned repeated crossing steps may be defined as comprising sufficient inbreeding to produce the inbred line. In the method, it may be desirable to select particular plants resulting from step (c) for continued crossing according to steps (b) and (c). By selecting plants having one or more desirable traits, a plant derived from hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 is obtained which possesses some of the desirable traits of the line/hybrid as well as potentially other selected traits.
In certain embodiments, the present invention provides a method of producing food or feed comprising: (a) obtaining a plant of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874, wherein the plant has been cultivated to maturity, and (b) collecting at least one squash from the plant.
In still yet another aspect of the invention, the genetic complement of squash hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 is provided. The phrase “genetic complement” is used to refer to the aggregate of nucleotide sequences, the expression of which sequences defines the phenotype of, in the present case, a squash plant, or a cell or tissue of that plant. A genetic complement thus represents the genetic makeup of a cell, tissue or plant, and a hybrid genetic complement represents the genetic make up of a hybrid cell, tissue or plant. The invention thus provides squash plant cells that have a genetic complement in accordance with the squash plant cells disclosed herein, and seeds and plants containing such cells.
Plant genetic complements may be assessed by genetic marker profiles, and by the expression of phenotypic traits that are characteristic of the expression of the genetic complement, e.g., isozyme typing profiles. It is understood that hybrid SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 could be identified by any of the many well known techniques such as, for example, Simple Sequence Length Polymorphisms (SSLPs) (Williams et al., Nucleic Acids Res., 1 8:6531-6535, 1990), Randomly Amplified Polymorphic DNAs (RAPDs), DNA Amplification Fingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs), Arbitrary Primed Polymerase Chain Reaction (AP-PCR), Amplified Fragment Length Polymorphisms (AFLPs) (EP 534 858, specifically incorporated herein by reference in its entirety), and Single Nucleotide Polymorphisms (SNPs) (Wang et al., Science, 280:1077-1082, 1998).
In still yet another aspect, the present invention provides hybrid genetic complements, as represented by squash plant cells, tissues, plants, and seeds, formed by the combination of a haploid genetic complement of a squash plant of the invention with a haploid genetic complement of a second squash plant, preferably, another, distinct squash plant. In another aspect, the present invention provides a squash plant regenerated from a tissue culture that comprises a hybrid genetic complement of this invention.
Any embodiment discussed herein with respect to one aspect of the invention applies to other aspects of the invention as well, unless specifically noted.
The term “about” is used to indicate that a value includes the standard deviation of the mean for the device or method being employed to determine the value. The use of the term “or” in the claims is used to mean “and/or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive. When used in conjunction with the word “comprising” or other open language in the claims, the words “a” and “an” denote “one or more,” unless specifically noted otherwise. The terms “comprise,” “have” and “include” are open-ended linking verbs. Any forms or tenses of one or more of these verbs, such as “comprises,” “comprising,” “has,” “having,” “includes” and “including,” are also open-ended. For example, any method that “comprises,” “has” or “includes” one or more steps is not limited to possessing only those one or more steps and also covers other unlisted steps. Similarly, any plant that “comprises,” “has” or “includes” one or more traits is not limited to possessing only those one or more traits and covers other unlisted traits.
Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and any specific examples provided, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.


DETAILED DESCRIPTION OF THE INVENTION
The invention provides methods and compositions relating to plants, seeds and derivatives of squash hybrid SV0116YL, squash line ZGN-EH10004 and squash LEB-EH-08-874.
Squash hybrid SV0116YL is also known as LEBEHH0116. SV0116YL is a medium length Lebanese-type squash hybrid that features high yield potential, strong adaptation to cool conditions, and resistance to some viral and fungal pathogens. SV0116YL is adapted for open field and protected culture growing conditions.
A. Origin and Breeding History of Squash Hybrid SV0116YL
The parents of hybrid SV0116YL are ZGN-EH10004 and LEB-EH-08-874. These parents were created as follows:
Female parent LEB-EH-08-874 is shared with other hybrids including “Jumann” (04858026) disclosed in U.S. Ser. No. 13/283,470, ROULAH, and SV0153YL disclosed in U.S. Ser. No. 13/764,768), for example.
LEB-EH-08-874 was developed through a minimum of 21 generations of inbreeding (controlled self pollination) and crossing. The initial source of the key traits of value (resistance to Zucchini Yellow Mosaic Virus (ZYMV) and Watermelon Mosaic Virus II (WMV-II)) was a diverse early generation population of squash distributed by Henry Munger of Cornell University in 1986 to collaborating seed companies and initially designated as “HMZYR” (presumably representing Henry Munger ZYMV resistant family). An individual from that population was crossed to a selection from the landrace “Small Green Algerian” in 1986, and then self pollinated for 3 generations, with selection for fruit color and shape in each generation (generations 1-4). An individual was selected from this population after the 3 generations of inbreeding and crossed to a proprietary breeding line known as LEB 47-42 (derived from a selection from cultivated landraces of squash common throughout the Middle East region). This germplasm was selected and self- and/or sib-pollinated for seven generations (generations 5-11), when an individual plant was crossed again to LEB-47-42 (generation 12). The progeny of this cross was selfed for 2 generations (12-13), then crossed to the proprietary inbred breeding line (generation 14) known as LEB-47-98 (itself a selection from a cultivated landrace of squash common in the Middle East region). This cross was then self pollinated for a minimum of 5 generations (15-20) before designating the line as LEB-EH-08-874. In generation 21 following the original selections this line was verified as phenotypically uniform for the purposes of commercial hybrid seed production.
Line ZGN-EH10004 was developed through a pedigree breeding strategy, and the initial founding germplasm includes only three items: a proprietary green zucchini inbred line known as G42, a selection from the Italian Landrace known as “Verde Milano Migliorato,” and a germplasm accession known as “C88” that was distributed by INRA, the French federal agricultural research agency.
To begin, the germplasm accession C88 was crossed to Verde Milano Migliorato, and the F2 and F3 generations were selected for resistance to ZYMV and Powdery Mildew (Podosphaera xanthii). Individual F3 plants from families showing high levels of resistance were selected and backcrossed to Verde Milano Migliorato. This process was repeated twice to create BC2F3 individuals. Following two additional generations of self pollinations, a BC2F5 individual was selected for further breeding.
The uniform inbred line G42 was initially crossed to the BC2F5 individual described above, and the F1 and F2 generations were self pollinated. In the F3 generation, families were screened for resistance to ZYMV and PM. Among families showing multiple disease resistant plants, an individual plant was chosen and backcrossed to the same inbred line. This process was repeated 4 times (hybridization, two generations of self pollination, disease screening, backcross to the initial G42 inbred line) so that the founding population leading to ZGN-EH10004 represents a genotype very similar to the G42 inbred, but with the addition of some key resistance traits from the BC2F5 individual described above. After the fourth backcross, an individual line was self pollinated through single seed descent to the BC4F6 generation, and an individual plant was designated as ZGN-EH10004. All self pollinated progeny of this individual BC4F6 plant were bulked to create the inbred line for hybrid seed production and eventual commercialization.
The parent lines are uniform and stable, as is a hybrid produced therefrom. A small percentage of variants can occur within commercially acceptable limits for almost any characteristic during the course of repeated multiplication. However no variants are expected.
B. Physiological and Morphological Characteristics of Squash Hybrid SV0116YL, Squash Line ZGN-EH10004 and Squash LEB-EH-08-874
In accordance with one aspect of the present invention, there is provided a plant having the physiological and morphological characteristics of squash hybrid SV0116YL and the parent lines thereof. A description of the physiological and morphological characteristics of such plants is presented in Tables 1-3.







TABLE 1







Physiological and Morphological Characteristics of Hybrid SV0116YL













Comparison Variety



CHARACTERISTIC
SV0116YL
Anita














1.
Species
Pepo
Pepo


2.
Kind/Use
squash
squash


3.
Type
summer (vegetable marrow)
summer


4.
Cotyledon













length
47.7
mm
42.55
mm



width
31
mm
28.55
mm











apex
rounded
tapered



veining
plainly visible
plainly visible



color
medium green
medium green



color (RHS Color Chart)
137B
137A



Seedling



shape of cotyledons
narrow elliptic (Bianchini)
elliptic



intensity of green color of
medium (Cora)
medium



cotyledons



cross section of
straight (Sunburst)
straight



cotyledons


5.
Mature Plant



growth habit
bush
semi-bush



plant type
glabrous
pilose


6.
Main Stem



cross-section shape
angled
round













diameter at mid-point of
27.8
mm
26.15
mm



1st internode



average length
33
cm
45.8
cm











average number of
  33.2
 33



internodes



Stem



color
completely green (Becky)
partly green and partly





yellow



intensity of green color
dark (Greyzini)
very dark



mottling
absent (Cinderella)
absent



tendrils
absent to rudimentary
well developed




(Goldrush, Sylvana)



Plant



growth habit
bush (Greyzini)
semi-trailing



branching
absent (Goldi)
absent



bush varieties only:
erect (Blanche non coureuse



attitude of petiole
3, Yellow Crookneck)



(excluding lower external



leaves)


7.
Leaves



blade shape
reniform
reniform



blade form
deep lobed
deep lobed



margin
dentate
denticulate



margin edges
frilled
frilled













average width
34.5
cm
36.2
cm



average length
27.6
cm
29.75
cm











leaf surface
blistered
blistered



dorsal surface pubescence
bristled
glabrous



vental surface pubescence
bristled
glabrous



color
dark green
dark green



color (RHS Color Chart)
147A
147A



leaf blotching
blotched with gray
blotched with gray



leaf blade: size
medium (Ambassador)
large



leaf blade: incisions
medium (Jackpot)
medium



leaf blade: intensity of
dark (Everest)
dark



green color of upper



surface



leaf blade: silvery patches
present (Civac)
present



leaf blade: relative area
small (Aziz)
small



covered by silvery patches













average petiole length
34
cm
38.35
cm











petiole length
medium (Goldi)
long



petiole: number of
very many (Yellow Summer
few



prickles
Crookneck)


8.
Flower













pistillate flower: average
19
cm
15.9
cm



diameter











pistillate flower: ovary
drum-like
drum-like













pistillate flower: average
1.4
cm
1.7
cm



pedicel length











pistillate flower: margin
curved
curved



shape



pistillate flower: margin
frilled
frilled



edges













pistillate flower: average
1.8
mm
1.35
mm



sepal width



pistillate flower: average
5.4
mm
5.4
mm



sepal length











pistillate flower: color
orange
orange



pistillate flower: color
 23A
 23A



(RHS Color Chart)













staminate flower: average
17.9
mm
16.8
mm



sepal length



staminate flower: average
4
mm
3
mm



sepal width



staminate flower: average
95.4
mm
122.9
mm



pedicel length











staminate flower: color
orange
orange



female flower: ring at
present (Aurore)
present



inner side of corolla



female flower: color of
green (Aurore, Early White
green



ring at inner side of
Bush Scallop, President)



corolla



female flower: intensity of
medium (Samba, Senator)
medium



green color of ring at inner



side of corolla (varieties



with green ring at inner



side of corolla)



male flower: ring at inner
present (Goldi)
present



side of corolla



male flower: color of ring
green (Austral, Belor, Goldi)
green



at inner side of corolla



male flower: intensity of
strong (Goldi)
medium



green color of ring at inner



side of corolla



staminate flower: color
 24A
 17A


9.
Fruit













market maturity: average
14.2
cm
13.3
cm



length



market maturity: average
2.4
cm
3.1
cm



width - stem end



at market maturity:
1.5
cm
3
cm



average width - blossom



end



market maturity: average
254.4
gm
236.5
gm



weight











market maturity: shape
straightneck
straightneck



according to variety type



market maturity: apex
rounded
flattened



market maturity: base
rounded
rounded



market maturity: ribs
inconspicuous
inconspicuous



market maturity: rib
shallow
shallow



furrow depth



market maturity: rib
medium wide
narrow



furrow width



market maturity: fruit
smooth
smooth



surface



market maturity: warts
none
none



market maturity: blossom
raised acorn
raised acorn



scar button



young fruit: ratio length/
medium (Cora)
medium



maximum diameter



(zucchini type varieties)



young fruit: general shape
cylindrical (Ambassador,
tapered elliptical



(zucchini and rounded
Ibis)



zucchini type varieties)



young fruit: main color of
green (Elite, Opal, Romano)
green



skin (excluding color of



ribs or grooves)



young fruit: intensity of
very light (Clarita, Goya,
light



green color of skin (only
Patty Green Tint)



varieties with green color



of skin)



general shape
cylindrical
club shaped



length (zucchini type
medium (Cora)
medium



varieties)



maximum diameter
medium (Opal)
medium



(zucchini type varieties)



ratio length/maximum
medium (Cora)
medium



diameter (zucchini type



varieties)



blossom end (zucchini and
rounded
rounded



neck type varieties)



grooves
absent
absent



ribs
present
present



protrusion of ribs
very weak (Leda, Tivoli)
weak



main color of skin
green (Ambassador, Baby
green



(excluding color of dots,
Bear)



patches, stripes and bands)



intensity of green color of
very light
light



skin (only varieties with



green color of skin)



color of ribs compared to
same (Grey Zucchini)
same



main color of skin



dots
present (Gold Rush, Table
present




Queen)



size of main dots
medium (Grey Zucchini)
small



secondary green color
absent (Grey Zucchini,
absent



between ribs (excluding
Small Sugar)



dots)



warts on skin
absent
absent



size of flower scar
medium (Spidi)
large



length of peduncle
medium (Cinderella)
medium



color of peduncle
green (Ambassador)
green



intensity of green color of
medium (Sunburst)
medium



peduncle



mottling of peduncle
present (Elite)
absent



ripe fruit: secondary color
yellow (Gold Rush)
yellow



of skin (excluding color of



mottles, patches, stripes



and bands)



ripe fruit: intensity of
medium
light



main color of skin (only



yellow and orange)



ripe fruit: color of flesh
cream (Elite)
cream



ripe fruit: lignified rind
present (Elite, Little Gem,
present




Scallopini, Yellow Summer




Crookneck)



ripe fruit: structure of
fibrous (Vegetable
fibrous



flesh
Spaghetti)


10.
Rind













average thickness at
2.8
mm
2.5
mm



medial











toughness
hard
hard



overall color pattern
regular
regular



main or ground color
yellow
yellowish-orange



main or ground color
 13A
 18A



(RHS Color Chart)


11.
Flesh













average blossom end
54.1
mm
29.45
mm



thickness



average medial thickness
55.1
mm
52.8
mm



average stem end
45.2
mm
26.25
mm



thickness











texture (fine, granular,
fine
fine



lumpy or stringy)



texture (soft, firm or
soft
firm



brittle)



texture (dry, moist or
juicy
juicy



juicy)



flavor
slightly sweet
sweet



quality
good
excellent



color
creamy-white
greenish-white



color (RHS Color Chart)
150D
157D


12.
Seed Cavity













average length
29.5
cm
25.5
cm



average width
7.1
cm
8.7
cm











location
conforms to fruit shape
conforms to fruit shape



placental tissue
abundant
abundant



center core
inconspicuous
inconspicuous


13.
Fruit Stalks













average length
2.2
cm
3.15
cm



average diameter
2
cm
2.1
cm











cross-section shape
irregular
irregular



twisting
twisted
not twisted



tapering
tapered
tapered



straightness
slightly curved
straight



texture
spongy
soft



furrows
shallow
shallow



surface
rough
spiny



attachment end
slightly expanded
expanded



detaches
with difficulty
easily



color
dark green
medium green



color (RHS Color Chart)
138A
143B


14.
Seeds













average length
17
mm
18.6
mm



average width
9.9
mm
9.2
mm



average thickness
2.2
mm
2.8
mm











face surface
wrinkled
smooth



color
cream
white



color (RHS Color Chart)
 18D
155A



luster
dull
dull



margin
straight
curved



margin edge
wedge-like
rounded



separation from pulp
moderately easy
easy













average grams per 100
13
gm
17
gm



seeds











average number of seeds
456
342



per fruit



seed coat
normal
normal



size
medium (Diamant)
large



shape
broad elliptic (Baby Boo)
elliptic



hull
present (Baby Bear, Elite)
present



appearance of hull
fully developed (Elite)
fully developed



color of hull
cream (De Nice à Fruit
cream




Rond)



fruit type:
zucchini



fruit: patches, stripes or
absent (Ambassador, Black



bands in ripe stage (if
Jack)



zucchini type)







*These are typical values. Values may vary due to environment. Other values that are substantially equivalent are also within the scope of the invention.













TABLE 2







Physiological and Morphological Characteristics of Line ZGN-EH10004













Comparison Variety



CHARACTERISTIC
ZGN-EH10004
Payroll














1.
Species
Pepo
Pepo


2.
Kind/Use
squash
squash


3.
Type
summer (vegetable marrow)
summer


4.
Cotyledon













length
43.3
mm
45.5
mm



width
26.4
mm
29.25
mm











apex
rounded
tapered



veining
prominent
obscure



color
dark green
medium green



color (RHS Color Chart)
137A
137A



Seedling



shape of cotyledons
narrow elliptic (Bianchini)
elliptic



intensity of green color of
dark (Lidia)
medium



cotyledons



cross section of
straight (Sunburst)
concave



cotyledons


5.
Mature Plant



growth habit
bush
bush



plant type
glabrous
pilose


6.
Main Stem



cross-section shape
angled
round













diameter at mid-point of
25.9
mm
23.4
mm



1st internode



average length
24
cm
30.75
cm











average number of
  30.6
  24.2



internodes



Stem



color
completely green (Becky)
completely green



intensity of green color
dark (Greyzini)
dark



mottling
absent (Cinderella)
absent



tendrils
absent to rudimentary
well developed




(Goldrush, Sylvana)



Plant



growth habit
bush (Greyzini)
bush



branching
absent (Goldi)
absent



degree of branching



bush varieties only:
erect to semi-erect (Sardane)
erect



attitude of petiole



(excluding lower external



leaves)


7.
Leaves



blade shape
reniform
reniform



blade form
deep lobed
deep lobed



margin
denticulate
dentate



margin edges
frilled
frilled













average width
36.5
cm
32.4
cm



average length
26.3
cm
29.1
cm











leaf surface
blistered
blistered



dorsal surface pubescence
soft hairy
soft hairy



vental surface pubescence
soft hairy
soft hairy



color
dark green
dark green



color (RHS Color Chart)
147A
139A



leaf blotching
blotched with gray
blotched with gray



leaf blade: size
medium (Ambassador)
small



leaf blade: incisions
medium (Jackpot)
deep



leaf blade: intensity of
dark (Everest)
dark



green color of upper



surface



leaf blade: silvery patches
present (Civac)
present



leaf blade: relative area
medium (Ambassador)
small



covered by silvery patches













average petiole length
34.6
cm
35.15
cm











petiole length
medium (Goldi)
long



petiole: number of
few (Opaline)
few



prickles


8.
Flower













pistillate flower: average
17.4
cm
15.3
cm



diameter











pistillate flower: ovary
drum-like
drum-like













pistillate flower: average
1.2
cm
1.75
cm



pedicel length











pistillate flower: margin
curved
straight



shape



pistillate flower: margin
frilled
frilled



edges













pistillate flower: average
1.2
mm
1.25
mm



sepal width



pistillate flower: average
4.4
mm
5.25
mm



sepal length











pistillate flower: color
orange
orange



pistillate flower: color
 23A
 17A



(RHS Color Chart)













staminate flower: average
13.5
mm
18.5
mm



sepal length



staminate flower: average
3.2
mm
2.95
mm



sepal width



staminate flower: average
61.2
mm
175
mm



pedicel length











staminate flower: color
orange
orange



female flower: ring at
present (Aurore)
present



inner side of corolla



female flower: color of
green (Aurore, Early White
green



ring at inner side of
Bush Scallop, President)



corolla



female flower: intensity of
medium (Samba, Senator)
medium



green color of ring at inner



side of corolla (varieties



with green ring at inner



side of corolla)



male flower: ring at inner
present (Goldi)
present



side of corolla



male flower: color of ring
green (Austral, Belor, Goldi)
green



at inner side of corolla



male flower: intensity of
strong (Goldi)
strong



green color of ring at inner



side of corolla



staminate flower: color
 23A
 21A


9.
Fruit













market maturity: average
16.1
cm
19.05
cm



length



market maturity: average
2.1
cm
3.2
cm



width - stem end



at market maturity:
0.9
cm
1.8
cm



average width - blossom



end



market maturity: average
193
gm
239
gm



weight











market maturity: shape
straightneck
straightneck



according to variety type



market maturity: apex
rounded
rounded



market maturity: base
rounded
taper pointed



market maturity: ribs
none
prominent



market maturity: fruit
smooth
smooth



surface



market maturity: warts
none
none



market maturity: blossom
slightly extended
raised acorn



scar button



young fruit: ratio length/
small (Opal)
large



maximum diameter



(zucchini type varieties)



young fruit: general shape
tapered elliptical (Top Kapi)
cylindrical



(zucchini and rounded



zucchini type varieties)



young fruit: main color of
green (Elite, Opal, Romano)
green



skin (excluding color of



ribs or grooves)



young fruit: intensity of
dark (Arlesa, Sandra, Zefira)
medium



green color of skin



(excluding color of ribs or



grooves; only varieties



with green color of skin)



general shape
cylindrical
cylindrical



length (zucchini type
medium (Cora)
very long



varieties)



maximum diameter
medium (Opal)
medium



(zucchini type varieties)



ratio length/maximum
large (Carlotta)
large



diameter (zucchini type



varieties)



blossom end (zucchini and
rounded
pointed



neck type varieties)



grooves
absent
absent



ribs
absent
present



main color of skin
green (Ambassador, Baby
green



(excluding color of dots,
Bear)



patches, stripes and bands)



intensity of green color of
dark (Cora)
medium



skin (only varieties with



green color of skin)



dots
present (Gold Rush, Table
present




Queen)



size of main dots
very small (Badger Cross)
small



warts on skin
absent
absent



size of flower scar
small (Goldi)
small



length of peduncle
very short (Arlesa)
short



color of peduncle
green (Ambassador)
green



intensity of green color of
medium (Sunburst)
medium



peduncle



mottling of peduncle
present (Elite)
absent



ripe fruit: main color of
cream (Bianchini, Opal)
yellow



skin (excluding color of



mottles, patches, stripes



and bands)



ripe fruit: secondary color
green
orange



of skin (excluding color of



mottles, patches, stripes



and bands)



green hue (only white and
present (Amalthee)



cream)



ripe fruit: prominence of
strong (Amalthee)



green hue (only white and



cream)



ripe fruit: color of flesh
cream (Elite)
cream



ripe fruit: lignified rind
present (Elite, Little Gem,
present




Scallopini, Yellow Summer




Crookneck)



ripe fruit: structure of
not fibrous (Elite)
fibrous



flesh


10.
Rind













average thickness at
1.6
mm
1.9
mm



medial











toughness
hard
hard



overall color pattern
regular
regular



main or ground color
green
bronze



main or ground color
N189A  
139A



(RHS Color Chart)



color of blotches
orange



color of blotches (RHS
 23A



Color Chart)



pattern of blotches
not specific


11.
Flesh













average blossom end
36.8
mm
22
mm



thickness



average medial thickness
41.6
mm
59
mm



average stem end
42.4
mm
27.4
mm



thickness











texture (fine, granular,
fine
fine



lumpy or stringy)



texture (soft, firm or
soft
soft



brittle)



texture (dry, moist or
moist
moist



juicy)



flavor
insipid
slightly sweet



quality
good
good



color
whitish cream
whitish cream



color (RHS Color Chart)
 4D
151B


12.
Seed Cavity













average length
22.7
cm
28.85
cm



average width
3.5
cm
5
cm











location
conforms to fruit shape
conforms to fruit shape



placental tissue
abundant
abundant



center core
inconspicuous
inconspicuous


13.
Fruit Stalks













average length
1.3
cm
2.75
cm



average diameter
1.4
cm
2.05
cm











cross-section shape
round
irregular



twisting
twisted
twisted



tapering
tapered
tapered



straightness
slightly curved
curved



texture
hard
soft



furrows
none
deep



surface
rough
rough



attachment end
slightly expanded
slightly expanded



detaches
with difficulty
easily



color
dark green
medium green



color (RHS Color Chart)
139A
137B


14.
Seeds













average length
13.2
mm
14.4
mm



average width
8.5
mm
8.5
mm



average thickness
2.1
mm
2.75
mm











face surface
smooth
smooth



color
white
white



color (RHS Color Chart)
155A
155A



luster
glossy
dull



margin
curved
curved



margin edge
rounded
rounded



separation from pulp
moderately easy
easy













average grams per 100
9.1
gm
13.75
gm



seeds











average number of seeds
63
187



per fruit



seed coat
normal
normal



size
small (Delicata)
small



shape
broad elliptic (Baby Boo)
elliptic



hull
present (Baby Bear, Elite)
present



appearance of hull
fully developed (Elite)
fully developed



color of hull
whitish (Table Queen)
cream



fruit type:
zucchini



fruit: patches, stripes or
present (Elite, Greyzini)



bands in ripe stage (if



zucchini type)







*These are typical values. Values may vary due to environment. Other values that are substantially equivalent are also within the scope of the invention.













TABLE 3







Physiological and Morphological Characteristics of LEB-EH-08-874













Comparison Variety



Characteristic
LEB-EH-08-874
Anita














1.
Species
Pepo
Pepo


2.
Kind/Use
squash
squash


3.
Type
summer (vegetable
summer




marrow)


4.
Cotyledon













length
41.05
mm
51.9
mm



width
33.4
mm
30.5
mm











apex
rounded
rounded



veining
obscure
plainly visible



color
medium green
light green



color (RHS Color Chart)
137A
137B



Seedling



shape of cotyledons
broad elliptic
elliptic




(Cinderella, Goldi)



intensity of green color of
medium (Cora)
light



cotyledons



cross section of cotyledons
concave
straight


5.
Mature Plant



growth habit
bush
bush



plant type
prickly
prickly


6.
Main Stem



cross-section shape
round
round













average diameter at mid-point
31.6
mm
29.2
mm



of 1st internode



average length
38.25
cm
31.6
cm











average number of internodes
 20.6
 23.5



Stem



color
completely green
partly green and partly




(Becky)
yellow



intensity of green color
medium (Cinderella)
medium



mottling
present (Cora)
present



tendrils
absent to rudimentary
absent to rudimentary




(Goldrush, Sylvana)



Plant



growth habit
bush (Greyzini)
bush



branching
absent (Goldi)
absent



bush varieties only: attitude of
semi-erect to horizontal
semi-erect



petiole (excluding lower
(Goldi)



external leaves)


7.
Leaves



blade shape
reniform
reniform



blade form
shallow lobed
deep lobed



margin
entire
denticulate



margin edges
frilled
frilled













average width
36.95
cm
36.8
cm



average length
27.55
cm
29
cm











leaf surface
blistered
blistered



dorsal surface pubescence
soft hairy
soft hairy



vental surface pubescence
soft hairy
soft hairy



color
dark green
dark green



color (RHS Color Chart)
147A
139A



leaf blotching
not blotched
blotched with gray



leaf blade: size
medium (Ambassador)
large



leaf blade: incisions
shallow (Everest)
medium



leaf blade: intensity of green
dark (Everest)
dark



color of upper surface



leaf blade: silvery patches
absent (Black Forest,
present




Scallopini)













average petiole length
34.6
cm
33.05
cm











petiole length
short (Jack Be Little,
medium




Karioka)



petiole: number of prickles
few (Opaline)
medium


8.
Flower













pistillate flower: diameter
14.4
cm
13.7
cm











pistillate flower: ovary
drum-like
drum-like













pistillate flower: average
1.7
cm
2
cm



pedicel length











pistillate flower: margin shape
curved
curved



pistillate flower: margin edges
frilled
plain













pistillate flower: average sepal
1.55
mm
1.5
mm



width



pistillate flower: average sepal
5.7
mm
6.5
mm



length











pistillate flower: color
orange
deep yellow



pistillate flower: color (RHS
 23A
 17A



Color Chart)













staminate flower: average
10.8
mm
16.3
mm



sepal length



staminate flower: average
2.55
mm
2.5
mm



sepal width



staminate flower: average
156.13
mm
147
mm



pedicel length











staminate flower: color
orange
deep yellow



female flower: ring at inner
present (Aurore)
present



side of corolla



female flower: color of ring at
green (Aurore, Early
green



inner side of corolla
White Bush Scallop,




President)



female flower: intensity of
weak (Maya, Sardane)
medium



green color of ring at inner



side of corolla (varieties with



green ring at inner side of



corolla)



male flower: ring at inner side
present (Goldi)
present



of corolla



male flower: color of ring at
green (Austral, Belor,
green



inner side of corolla
Goldi)



male flower: intensity of
medium (Verdi)
medium



green color of ring at inner



side of corolla (varieties with



green ring at inner side of



corolla)



staminate flower: color
 23A
 21A


9.
Fruit













market maturity: average
10.8
cm
12.4
cm



length



market maturity: average
4.87
cm
3.8
cm



width - stem end



market maturity: average
5.4
cm
4.6
cm



width - blossom end



market maturity: average
230.7
gm
208.4
gm



weight











market maturity: shape
banana
straightneck



according to variety type



market maturity: apex
rounded
taper pointed



market maturity: base
flattened
flattened



market maturity: ribs
inconspicuous
inconspicuous



market maturity: rib furrow
narrow
narrow



width



market maturity: fruit surface
smooth
smooth



market maturity: warts
none
none



market maturity: blossom scar
raised acorn
raised acorn



button



young fruit: ratio length/
small (Opal)
small



maximum diameter (zucchini



type varieties)



young fruit: general shape
pear shaped (Clarita)
pear shaped



(zucchini and rounded



zucchini type varieties)



young fruit: main color of
green (Elite, Opal,
partly white and partly



skin (excluding color of ribs
Romano)
green



or grooves)



young fruit: intensity of green
light (Arlika)
light



color of skin (excluding color



of ribs or grooves; only



varieties with green color of



skin)



general shape
pear shaped
pear shaped



length (zucchini type
short (Jedida)
short



varieties)



maximum diameter (zucchini
large (Jericho, Spidy)
large



type varieties)



ratio length/maximum
large (Carlotta)
medium



diameter (zucchini type



varieties)



blossom end (zucchini and
rounded
rounded



neck type varieties)



grooves
absent
absent



ribs
present
present



protrusion of ribs
weak (Ambassador)
weak



main color of skin (excluding
green (Ambassador,
green



color of dots, patches, stripes
Baby Bear)



and bands)



intensity of green color of
light
very light



skin (excluding color of dots,



patches, stripes and bands;



varieties with green color or



skin)



stripes in grooves
absent (Baby Bear,
absent




Jack Be Little)



color of ribs compared to
same (Grey Zucchini)
same



main color of skin (excluding



color of dots, patches, stripes



and bands)



dots
present (Gold Rush,
present




Table Queen)



size of main dots
medium (Grey
large




Zucchini)



secondary green color
absent (Grey Zucchini,



between ribs (excluding dots)
Small Sugar)



warts on skin
absent
absent



size of flower scar
large (Cinderella)
medium



length of peduncle
medium (Cinderella)
long



color of peduncle
green (Ambassador)
green



intensity of green color of
medium (Sunburst)
medium



peduncle



mottling of peduncle
absent (Sunburst)
present



type
rounded zucchini



patches, stripes or bands in
absent (Ambassador,



ripe stage
Black Jack)



ripe fruit: main color of skin
yellow (Gold Rush)
yellow



(excluding color of mottles,



patches, stripes and bands)



ripe fruit: intensity of main
light
dark



color of skin (only yellow and



orange)



ripe fruit: color of flesh
cream (Elite)
yellow



ripe fruit: lignified rind
present (Elite, Little
present




Gem, Scallopini,




Yellow Summer




Crookneck)



ripe fruit: structure of flesh
fibrous (Vegetable
fibrous




Spaghetti)


10.
Rind













average thickness at medial
4.2
mm
3.1
mm











toughness
hard
hard



overall color pattern
regular
irregular



main or ground color
yellow
creamy-yellow



main or ground color (RHS
160B
 20C



Color Chart)


11.
Flesh













average blossom end
18.66
mm
14.4
mm



thickness



average medial thickness
85.4
mm
78.9
mm



average stem end thickness
18.9
mm
17.6
mm











texture (fine, granular, lumpy
fine
stringy



or stringy)



texture (soft, firm or brittle)
firm
firm



texture (dry, moist or juicy)
juicy
moist



flavor
sweet
insipid



quality
excellent
good



color
off-white
cream



color (RHS Color Chart)
155C
155C


12.
Seed Cavity













average length
18.3
cm
28.1
cm



average width
10.18
cm
10.1
cm











location
conforms to fruit shape
conforms to fruit shape



placental tissues
abundant
abundant



center core
inconspicuous
prominent


13.
Fruit Stalks













average length
2.52
cm
3.3
cm



average diameter
2.39
cm
2.5
cm











cross-section shape
irregular
irregular



twisting
not twisted
not twisted



tapering
not tapered
not tapered



straightness
straight
straight



texture
hard
spongy



furrows
shallow
deep



surface
spiny
spiny



attachment end
expanded
not expanded



detaches
easily
easily



color
light green
medium green



color (RHS Color Chart)
144B
144B


14.
Seeds













average length
17.3
mm
16.3
mm



average width
10.11
mm
9.15
mm



average thickness
2.76
mm
2.5
mm











face surface
wrinkled
smooth



color
cream
cream



color (RHS Color Chart)
160D
162C



luster
dull
dull



margin
curved
straight



margin edge
rounded
rounded



separation from pulp
easy
easy













average grams per 100 seeds
19.1
gm
16.5
gm











average number of seeds per
174.5
346.5



fruit



seed coat
normal
normal



size
large
large



shape
elliptic (Elite)
broad elliptic



hull
present (Baby Bear,
present




Elite)



appearance of hull
fully developed (Elite)
rudimentary



color of hull
cream (De Nice à Fruit
cream




Rond)







*These are typical values. Values may vary due to environment. Other values that are substantially equivalent are also within the scope of the invention.






C. Breeding Squash Plants
One aspect of the current invention concerns methods for producing seed of squash hybrid SV0116YL involving crossing squash lines ZGN-EH10004 and LEB-EH-08-874. Alternatively, in other embodiments of the invention, hybrid SV0116YL, line ZGN-EH10004, or LEB-EH-08-874 may be crossed with itself or with any second plant. Such methods can be used for propagation of hybrid SV0116YL and/or the squash lines ZGN-EH10004 and LEB-EH-08-874, or can be used to produce plants that are derived from hybrid SV0116YL and/or the squash lines ZGN-EH10004 and LEB-EH-08-874. Plants derived from hybrid SV0116YL and/or the squash lines ZGN-EH10004 and LEB-EH-08-874 may be used, in certain embodiments, for the development of new squash varieties.
The development of new varieties using one or more starting varieties is well known in the art. In accordance with the invention, novel varieties may be created by crossing hybrid SV0116YL followed by multiple generations of breeding according to such well known methods. New varieties may be created by crossing with any second plant. In selecting such a second plant to cross for the purpose of developing novel lines, it may be desired to choose those plants which either themselves exhibit one or more selected desirable characteristics or which exhibit the desired characteristic(s) when in hybrid combination. Once initial crosses have been made, inbreeding and selection take place to produce new varieties. For development of a uniform line, often five or more generations of selfing and selection are involved.
Uniform lines of new varieties may also be developed by way of double-haploids. This technique allows the creation of true breeding lines without the need for multiple generations of selfing and selection. In this manner true breeding lines can be produced in as little as one generation. Haploid embryos may be produced from microspores, pollen, anther cultures, or ovary cultures. The haploid embryos may then be doubled autonomously, or by chemical treatments (e.g. colchicine treatment). Alternatively, haploid embryos may be grown into haploid plants and treated to induce chromosome doubling. In either case, fertile homozygous plants are obtained. In accordance with the invention, any of such techniques may be used in connection with a plant of the invention and progeny thereof to achieve a homozygous line.
Backcrossing can also be used to improve an inbred plant. Backcrossing transfers a specific desirable trait from one inbred or non-inbred source to an inbred that lacks that trait. This can be accomplished, for example, by first crossing a superior inbred (A) (recurrent parent) to a donor inbred (non-recurrent parent), which carries the appropriate locus or loci for the trait in question. The progeny of this cross are then mated back to the superior recurrent parent (A) followed by selection in the resultant progeny for the desired trait to be transferred from the non-recurrent parent. After five or more backcross generations with selection for the desired trait, the progeny have the characteristic being transferred, but are like the superior parent for most or almost all other loci. The last backcross generation would be selfed to give pure breeding progeny for the trait being transferred.
The plants of the present invention are particularly well suited for the development of new lines based on the elite nature of the genetic background of the plants. In selecting a second plant to cross with SV0116YL and/or squash lines ZGN-EH10004 and LEB-EH-08-874 for the purpose of developing novel squash lines, it will typically be preferred to choose those plants which either themselves exhibit one or more selected desirable characteristics or which exhibit the desired characteristic(s) when in hybrid combination. Examples of desirable traits may include, in specific embodiments, high seed yield, high seed germination, seedling vigor, high fruit yield, disease tolerance or resistance, and adaptability for soil and climate conditions. Consumer-driven traits, such as a fruit shape, color, texture, and taste are other examples of traits that may be incorporated into new lines of squash plants developed by this invention.
D. Performance Characteristics
As described above, hybrid SV0116YL exhibits desirable traits, as conferred by squash lines ZGN-EH10004 and LEB-EH-08-874. The performance characteristics of hybrid SV0116YL and squash lines ZGN-EH10004 and LEB-EH-08-874 were the subject of an objective analysis of the performance traits relative to other varieties. The results of the analysis are presented below.







TABLE 4







Performance Data from early summer 2012, near Amman Jordan. Ratings are on a


1-9 scale, where 1 is ideal, 5 is typical, and 9 is unacceptable or fully susceptible.


















Fruit
Fruit
Fruit
Blossom
Ease of
Viral



Marketable


Hybrid
shape
color
uniformity
scar
harvest
severity
Vigor
Habit
Spines
fruit/plant





SV0116YL
4
5
4
6
1
7
5
5
7
6.9


SV0116YL
3
5
4
8
1
4
4
5
8
6.9


SV0116YL
3
3
3
6
1
5
6
4
8
7.3


SV0116YL
5
5
3
8
1
4
4
4
7
7.3


SV0116YL
4
4
3
7
1
6
4
6
8
7.5


Anita
3
4
6
7
4
9
6
5
8
3.3


Anita
3
4
5
7
5
9
4
6
7
3.7


Anita
5
4
6
7
3
7
4
5
7
4.2


Anita
3
4
4
6
2
8
4
6
8
4.3


Anita
4
4
5
7
3
7
4
4
7
5.0









E. Further Embodiments of the Invention
In certain aspects of the invention, plants described herein are provided modified to include at least a first desired heritable trait. Such plants may, in one embodiment, be developed by a plant breeding technique called backcrossing, wherein essentially all of the morphological and physiological characteristics of a variety are recovered in addition to a genetic locus transferred into the plant via the backcrossing technique. The term single locus converted plant as used herein refers to those squash plants which are developed by a plant breeding technique called backcrossing, wherein essentially all of the morphological and physiological characteristics of a variety are recovered in addition to the single locus transferred into the variety via the backcrossing technique. By essentially all of the morphological and physiological characteristics, it is meant that the characteristics of a plant are recovered that are otherwise present when compared in the same environment, other than an occasional variant trait that might arise during backcrossing or direct introduction of a transgene.
Backcrossing methods can be used with the present invention to improve or introduce a characteristic into the present variety. The parental squash plant which contributes the locus for the desired characteristic is termed the nonrecurrent or donor parent. This terminology refers to the fact that the nonrecurrent parent is used one time in the backcross protocol and therefore does not recur. The parental squash plant to which the locus or loci from the nonrecurrent parent are transferred is known as the recurrent parent as it is used for several rounds in the backcrossing protocol.
In a typical backcross protocol, the original variety of interest (recurrent parent) is crossed to a second variety (nonrecurrent parent) that carries the single locus of interest to be transferred. The resulting progeny from this cross are then crossed again to the recurrent parent and the process is repeated until a squash plant is obtained wherein essentially all of the morphological and physiological characteristics of the recurrent parent are recovered in the converted plant, in addition to the single transferred locus from the nonrecurrent parent.
The selection of a suitable recurrent parent is an important step for a successful backcrossing procedure. The goal of a backcross protocol is to alter or substitute a single trait or characteristic in the original variety. To accomplish this, a single locus of the recurrent variety is modified or substituted with the desired locus from the nonrecurrent parent, while retaining essentially all of the rest of the desired genetic, and therefore the desired physiological and morphological constitution of the original variety. The choice of the particular nonrecurrent parent will depend on the purpose of the backcross; one of the major purposes is to add some commercially desirable trait to the plant. The exact backcrossing protocol will depend on the characteristic or trait being altered and the genetic distance between the recurrent and nonrecurrent parents. Although backcrossing methods are simplified when the characteristic being transferred is a dominant allele, a recessive allele, or an additive allele (between recessive and dominant), may also be transferred. In this instance it may be necessary to introduce a test of the progeny to determine if the desired characteristic has been successfully transferred.
In one embodiment, progeny squash plants of a backcross in which a plant described herein is the recurrent parent comprise (i) the desired trait from the non-recurrent parent and (ii) all of the physiological and morphological characteristics of squash the recurrent parent as determined at the 5% significance level when grown in the same environmental conditions.
New varieties can also be developed from more than two parents. The technique, known as modified backcrossing, uses different recurrent parents during the backcrossing. Modified backcrossing may be used to replace the original recurrent parent with a variety having certain more desirable characteristics or multiple parents may be used to obtain different desirable characteristics from each.
With the development of molecular markers associated with particular traits, it is possible to add additional traits into an established germ line, such as represented here, with the end result being substantially the same base germplasm with the addition of a new trait or traits. Molecular breeding, as described in Moose and Mumm, 2008 (Plant Physiology, 147: 969-977), for example, and elsewhere, provides a mechanism for integrating single or multiple traits or QTL into an elite line. This molecular breeding-facilitated movement of a trait or traits into an elite line may encompass incorporation of a particular genomic fragment associated with a particular trait of interest into the elite line by the mechanism of identification of the integrated genomic fragment with the use of flanking or associated marker assays. In the embodiment represented here, one, two, three or four genomic loci, for example, may be integrated into an elite line via this methodology. When this elite line containing the additional loci is further crossed with another parental elite line to produce hybrid offspring, it is possible to then incorporate at least eight separate additional loci into the hybrid. These additional loci may confer, for example, such traits as a disease resistance or a fruit quality trait. In one embodiment, each locus may confer a separate trait. In another embodiment, loci may need to be homozygous and exist in each parent line to confer a trait in the hybrid. In yet another embodiment, multiple loci may be combined to confer a single robust phenotype of a desired trait.
Many single locus traits have been identified that are not regularly selected for in the development of a new inbred but that can be improved by backcrossing techniques. Single locus traits may or may not be transgenic; examples of these traits include, but are not limited to, herbicide resistance, resistance to bacterial, fungal, or viral disease, insect resistance, modified fatty acid or carbohydrate metabolism, and altered nutritional quality. These comprise genes generally inherited through the nucleus.
Direct selection may be applied where the single locus acts as a dominant trait. For this selection process, the progeny of the initial cross are assayed for viral resistance and/or the presence of the corresponding gene prior to the backcrossing. Selection eliminates any plants that do not have the desired gene and resistance trait, and only those plants that have the trait are used in the subsequent backcross. This process is then repeated for all additional backcross generations.
Selection of squash plants for breeding is not necessarily dependent on the phenotype of a plant and instead can be based on genetic investigations. For example, one can utilize a suitable genetic marker which is closely genetically linked to a trait of interest. One of these markers can be used to identify the presence or absence of a trait in the offspring of a particular cross, and can be used in selection of progeny for continued breeding. This technique is commonly referred to as marker assisted selection. Any other type of genetic marker or other assay which is able to identify the relative presence or absence of a trait of interest in a plant can also be useful for breeding purposes. Procedures for marker assisted selection are well known in the art. Such methods will be of particular utility in the case of recessive traits and variable phenotypes, or where conventional assays may be more expensive, time consuming or otherwise disadvantageous. Types of genetic markers which could be used in accordance with the invention include, but are not necessarily limited to, Simple Sequence Length Polymorphisms (SSLPs) (Williams et al., Nucleic Acids Res., 1 8:6531-6535, 1990), Randomly Amplified Polymorphic DNAs (RAPDs), DNA Amplification Fingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs), Arbitrary Primed Polymerase Chain Reaction (AP-PCR), Amplified Fragment Length Polymorphisms (AFLPs) (EP 534 858, specifically incorporated herein by reference in its entirety), and Single Nucleotide Polymorphisms (SNPs) (Wang et al., Science, 280:1077-1082, 1998).
F. Plants Derived by Genetic Engineering
Many useful traits that can be introduced by backcrossing, as well as directly into a plant, are those which are introduced by genetic transformation techniques. Genetic transformation may therefore be used to insert a selected transgene into a plant of the invention or may, alternatively, be used for the preparation of transgenes which can be introduced by backcrossing. Methods for the transformation of plants that are well known to those of skill in the art and applicable to many crop species include, but are not limited to, electroporation, microprojectile bombardment, Agrobacterium-mediated transformation and direct DNA uptake by protoplasts.
To effect transformation by electroporation, one may employ either friable tissues, such as a suspension culture of cells or embryogenic callus or alternatively one may transform immature embryos or other organized tissue directly. In this technique, one would partially degrade the cell walls of the chosen cells by exposing them to pectin-degrading enzymes (pectolyases) or mechanically wound tissues in a controlled manner.
An efficient method for delivering transforming DNA segments to plant cells is microprojectile bombardment. In this method, particles are coated with nucleic acids and delivered into cells by a propelling force. Exemplary particles include those comprised of tungsten, platinum, and preferably, gold. For the bombardment, cells in suspension are concentrated on filters or solid culture medium. Alternatively, immature embryos or other target cells may be arranged on solid culture medium. The cells to be bombarded are positioned at an appropriate distance below the macroprojectile stopping plate.
An illustrative embodiment of a method for delivering DNA into plant cells by acceleration is the Biolistics Particle Delivery System, which can be used to propel particles coated with DNA or cells through a screen, such as a stainless steel or Nytex screen, onto a surface covered with target cells. The screen disperses the particles so that they are not delivered to the recipient cells in large aggregates. Microprojectile bombardment techniques are widely applicable, and may be used to transform virtually any plant species.
Agrobacterium-mediated transfer is another widely applicable system for introducing gene loci into plant cells. An advantage of the technique is that DNA can be introduced into whole plant tissues, thereby bypassing the need for regeneration of an intact plant from a protoplast. Modern Agrobacterium transformation vectors are capable of replication in E. coli as well as Agrobacterium, allowing for convenient manipulations (Klee et al., Bio-Technology, 3(7):637-642, 1985). Moreover, recent technological advances in vectors for Agrobacterium-mediated gene transfer have improved the arrangement of genes and restriction sites in the vectors to facilitate the construction of vectors capable of expressing various polypeptide coding genes. The vectors described have convenient multi-linker regions flanked by a promoter and a polyadenylation site for direct expression of inserted polypeptide coding genes. Additionally, Agrobacterium containing both armed and disarmed Ti genes can be used for transformation.
In those plant strains where Agrobacterium-mediated transformation is efficient, it is the method of choice because of the facile and defined nature of the gene locus transfer. The use of Agrobacterium-mediated plant integrating vectors to introduce DNA into plant cells is well known in the art (Fraley et al., Bio/Technology, 3:629-635, 1985; U.S. Pat. No. 5,563,055).
Transformation of plant protoplasts also can be achieved using methods based on calcium phosphate precipitation, polyethylene glycol treatment, electroporation, and combinations of these treatments (see, e.g., Potrykus et al., Mol. Gen. Genet., 199:183-188, 1985; Omirulleh et al., Plant Mol. Biol., 21(3):415-428, 1993; Fromm et al., Nature, 312:791-793, 1986; Uchimiya et al., Mol. Gen. Genet., 204:204, 1986; Marcotte et al., Nature, 335:454, 1988). Transformation of plants and expression of foreign genetic elements is exemplified in Choi et al. (Plant Cell Rep., 13: 344-348, 1994), and Ellul et al. (Theor. Appl. Genet., 107:462-469, 2003).
A number of promoters have utility for plant gene expression for any gene of interest including but not limited to selectable markers, scoreable markers, genes for pest tolerance, disease resistance, nutritional enhancements and any other gene of agronomic interest. Examples of constitutive promoters useful for plant gene expression include, but are not limited to, the cauliflower mosaic virus (CaMV) P-35S promoter, which confers constitutive, high-level expression in most plant tissues (see, e.g., Odel et al., Nature, 313:810, 1985), including in monocots (see, e.g., Dekeyser et al., Plant Cell, 2:591, 1990; Terada and Shimamoto, Mol. Gen. Genet., 220:389, 1990); a tandemly duplicated version of the CaMV 35S promoter, the enhanced 35S promoter (P-e35S); 1 the nopaline synthase promoter (An et al., Plant Physiol., 88:547, 1988); the octopine synthase promoter (Fromm et al., Plant Cell, 1:977, 1989); and the figwort mosaic virus (P-FMV) promoter as described in U.S. Pat. No. 5,378,619 and an enhanced version of the FMV promoter (P-eFMV) where the promoter sequence of P-FMV is duplicated in tandem; the cauliflower mosaic virus 19S promoter; a sugarcane bacilliform virus promoter; a commelina yellow mottle virus promoter; and other plant DNA virus promoters known to express in plant cells.
A variety of plant gene promoters that are regulated in response to environmental, hormonal, chemical, and/or developmental signals can also be used for expression of an operably linked gene in plant cells, including promoters regulated by (1) heat (Callis et al., Plant Physiol., 88:965, 1988), (2) light (e.g., pea rbcS-3A promoter, Kuhlemeier et al., Plant Cell, 1:471, 1989; maize rbcS promoter, Schaffner and Sheen, Plant Cell, 3:997, 1991; or chlorophyll a/b-binding protein promoter, Simpson et al., EMBO J., 4:2723, 1985), (3) hormones, such as abscisic acid (Marcotte et al., Plant Cell, 1:969, 1989), (4) wounding (e.g., wunl, Siebertz et al., Plant Cell, 1:961, 1989); or (5) chemicals such as methyl jasmonate, salicylic acid, or Safener. It may also be advantageous to employ organ-specific promoters (e.g., Roshal et al., EMBO J., 6:1155, 1987; Schernthaner et al., EMBO J., 7:1249, 1988; Bustos et al., Plant Cell, 1:839, 1989).
Exemplary nucleic acids which may be introduced to plants of this invention include, for example, DNA sequences or genes from another species, or even genes or sequences which originate with or are present in the same species, but are incorporated into recipient cells by genetic engineering methods rather than classical reproduction or breeding techniques. However, the term “exogenous” is also intended to refer to genes that are not normally present in the cell being transformed, or perhaps simply not present in the form, structure, etc., as found in the transforming DNA segment or gene, or genes which are normally present and that one desires to express in a manner that differs from the natural expression pattern, e.g., to over-express. Thus, the term “exogenous” gene or DNA is intended to refer to any gene or DNA segment that is introduced into a recipient cell, regardless of whether a similar gene may already be present in such a cell. The type of DNA included in the exogenous DNA can include DNA which is already present in the plant cell, DNA from another plant, DNA from a different organism, or a DNA generated externally, such as a DNA sequence containing an antisense message of a gene, or a DNA sequence encoding a synthetic or modified version of a gene.
Many hundreds if not thousands of different genes are known and could potentially be introduced into a squash plant according to the invention. Non-limiting examples of particular genes and corresponding phenotypes one may choose to introduce into a squash plant include one or more genes for insect tolerance, such as a Bacillus thuringiensis (B.t.) gene, pest tolerance such as genes for fungal disease control, herbicide tolerance such as genes conferring glyphosate tolerance, and genes for quality improvements such as yield, nutritional enhancements, environmental or stress tolerances, or any desirable changes in plant physiology, growth, development, morphology or plant product(s). For example, structural genes would include any gene that confers insect tolerance including but not limited to a Bacillus insect control protein gene as described in WO 99/31248, herein incorporated by reference in its entirety, U.S. Pat. No. 5,689,052, herein incorporated by reference in its entirety, U.S. Pat. Nos. 5,500,365 and 5,880,275, herein incorporated by reference in their entirety. In another embodiment, the structural gene can confer tolerance to the herbicide glyphosate as conferred by genes including, but not limited to Agrobacterium strain CP4 glyphosate resistant EPSPS gene (aroA:CP4) as described in U.S. Pat. No. 5,633,435, herein incorporated by reference in its entirety, or glyphosate oxidoreductase gene (GOX) as described in U.S. Pat. No. 5,463,175, herein incorporated by reference in its entirety.
Alternatively, the DNA coding sequences can affect these phenotypes by encoding a non-translatable RNA molecule that causes the targeted inhibition of expression of an endogenous gene, for example via antisense- or cosuppression-mediated mechanisms (see, for example, Bird et al., Biotech. Gen. Engin. Rev., 9:207, 1991). The RNA could also be a catalytic RNA molecule (i.e., a ribozyme) engineered to cleave a desired endogenous mRNA product (see for example, Gibson and Shillito, Mol. Biotech., 7:125, 1997). Thus, any gene which produces a protein or mRNA which expresses a phenotype or morphology change of interest is useful for the practice of the present invention.
G. Definitions
In the description and tables herein, a number of terms are used. In order to provide a clear and consistent understanding of the specification and claims, the following definitions are provided:
Allele: Any of one or more alternative forms of a gene locus, all of which alleles relate to one trait or characteristic. In a diploid cell or organism, the two alleles of a given gene occupy corresponding loci on a pair of homologous chromosomes.
Backcrossing: A process in which a breeder repeatedly crosses hybrid progeny, for example a first generation hybrid (F1), back to one of the parents of the hybrid progeny. Backcrossing can be used to introduce one or more single locus conversions from one genetic background into another.
Crossing: The mating of two parent plants.
Cross-pollination: Fertilization by the union of two gametes from different plants.
Diploid: A cell or organism having two sets of chromosomes.
Emasculate: The removal of plant male sex organs or the inactivation of the organs with a cytoplasmic or nuclear genetic factor or a chemical agent conferring male sterility.
Enzymes: Molecules which can act as catalysts in biological reactions.
F1 Hybrid: The first generation progeny of the cross of two nonisogenic plants.
Genotype: The genetic constitution of a cell or organism.
Haploid: A cell or organism having one set of the two sets of chromosomes in a diploid.
Linkage: A phenomenon wherein alleles on the same chromosome tend to segregate together more often than expected by chance if their transmission was independent.
Marker: A readily detectable phenotype, preferably inherited in codominant fashion (both alleles at a locus in a diploid heterozygote are readily detectable), with no environmental variance component, i.e., heritability of 1.
Phenotype: The detectable characteristics of a cell or organism, which characteristics are the manifestation of gene expression.
Quantitative Trait Loci (QTL): Quantitative trait loci (QTL) refer to genetic loci that control to some degree numerically representable traits that are usually continuously distributed.
Resistance: As used herein, the terms “resistance” and “tolerance” are used interchangeably to describe plants that show no symptoms to a specified biotic pest, pathogen, abiotic influence or environmental condition. These terms are also used to describe plants showing some symptoms but that are still able to produce marketable product with an acceptable yield. Some plants that are referred to as resistant or tolerant are only so in the sense that they may still produce a crop, even though the plants are stunted and the yield is reduced.
Regeneration: The development of a plant from tissue culture.
Royal Horticultural Society (RHS) color chart value: The RHS color chart is a standardized reference which allows accurate identification of any color. A color's designation on the chart describes its hue, brightness and saturation. A color is precisely named by the RHS color chart by identifying the group name, sheet number and letter, e.g., Yellow-Orange Group 19A or Red Group 41B.
Self-pollination: The transfer of pollen from the anther to the stigma of the same plant.
Single Locus Converted (Conversion) Plant: Plants which are developed by a plant breeding technique called backcrossing, wherein essentially all of the morphological and physiological characteristics of a squash variety are recovered in addition to the characteristics of the single locus transferred into the variety via the backcrossing technique and/or by genetic transformation.
Substantially Equivalent: A characteristic that, when compared, does not show a statistically significant difference (e.g., p=0.05) from the mean.
Tissue Culture: A composition comprising isolated cells of the same or a different type or a collection of such cells organized into parts of a plant.
Transgene: A genetic locus comprising a sequence which has been introduced into the genome of a squash plant by transformation.
H. Deposit Information
A deposit of squash hybrid SV0116YL and inbred parent lines ZGN-EH10004 and LEB-EH-08-874, disclosed above and recited in the claims, has been made with the American Type Culture Collection (ATCC), 10801 University Blvd., Manassas, Va. 20110-2209. The dates of deposits were Jul. 23, 2013, Jul. 23, 2013, and Sep. 23, 2011, respectively. The accession numbers for those deposited seeds of squash hybrid SV0116YL and inbred parent lines ZGN-EH10004 and LEB-EH-08-874 are ATCC Accession No. PTA-120512, ATCC Accession No. PTA-120511, and ATCC Accession No. PTA-12119, respectively. Upon issuance of a patent, all restrictions upon the deposits will be removed, and the deposits are intended to meet all of the requirements of 37 C.F.R. §1.801-1.809. The deposits will be maintained in the depository for a period of 30 years, or 5 years after the last request, or for the effective life of the patent, whichever is longer, and will be replaced if necessary during that period.
Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity and understanding, it will be obvious that certain changes and modifications may be practiced within the scope of the invention, as limited only by the scope of the appended claims.
All references cited herein are hereby expressly incorporated herein by reference.

## Claims

1. A squash plant comprising at least a first set of the chromosomes of squash line ZGN-EH10004, a sample of seed of said line having been deposited under ATCC Accession Number PTA-120511.

2. A seed comprising at least a first set of the chromosomes of squash line ZGN-EH10004, a sample of seed of said line having been deposited under ATCC Accession Number PTA-120511.

3. The plant of claim 1, which is an inbred.

4. The plant of claim 1, which is a hybrid.

5. The seed of claim 2, which is an inbred.

6. The seed of claim 2, which is a hybrid.

7. The plant of claim 4, wherein the hybrid plant is squash hybrid SV0116YL, a sample of seed of said hybrid SV0116YL having been deposited under ATCC Accession Number PTA-120512.

8. The seed of claim 6, defined as a seed of squash hybrid SV0116YL, a sample of seed of said hybrid SV0116YL having been deposited under ATCC Accession Number PTA-120512.

9. The seed of claim 2, defined as a seed of line ZGN-EH10004.

10. A plant part of the plant of claim 1, said plant part comprising at least a first set of the chromosomes of squash line ZGN-EH10004, a sample of seed of said line having been deposited under ATCC Accession Number PTA-120511.

11. The plant part of claim 10, further defined as a leaf, an ovule, pollen, a fruit, or a cell.

12. A squash plant having all the physiological and morphological characteristics of the squash plant of claim 7.

13. A tissue culture of regenerable cells of the plant of claim 1, said cells comprising at least a first set of the chromosomes of squash line ZGN-EH10004, a sample of seed of said line having been deposited under ATCC Accession Number PTA-120511.

14. The tissue culture according to claim 13, comprising cells or protoplasts from a plant part selected from the group consisting of embryos, meristems, cotyledons, pollen, leaves, anthers, roots, root tips, pistil, flower, seed and stalks.

15. A squash plant regenerated from the tissue culture of claim 13.

16. A method of vegetatively propagating the squash plant of claim 1, said method comprising the steps of:
(a) collecting tissue capable of being propagated from the plant according to claim 1;
(b) cultivating said tissue to obtain proliferated shoots; and
(c) rooting said proliferated shoots to obtain rooted plantlets.

17. The method of claim 16, further comprising growing at least a first squash plant from said rooted plantlets.

18. A method of introducing a desired trait into a squash line, said method comprising:
(a) crossing a plant of line ZGN-EH10004 with a second squash plant that comprises a desired trait to produce F1 progeny, a sample of seed of said line having been deposited under ATCC Accession Number PTA-120511;
(b) selecting an F1 progeny that comprises the desired trait;
(c) backcrossing the selected F1 progeny with a plant of line ZGN-EH10004 to produce backcross progeny;
(d) selecting backcross progeny comprising the desired trait; and
(e) repeating steps (c) and (d) three or more times to produce selected fourth or higher backcross progeny that comprise the desired trait and otherwise comprises all of the morphological and physiological characteristics of line ZGN-EH10004.

19. A method of producing a squash plant comprising an added trait, the method comprising introducing a transgene conferring the trait into a plant of squash hybrid SV0116YL, or squash line ZGN-EH10004, a sample of seed of said hybrid and line having been deposited under ATCC Accession Number PTA-120512, and ATCC Accession Number PTA-120511, respectively.

20. A plant of squash hybrid SV0116YL or squash line ZGN-EH10004, further comprising a transgene, wherein a sample of seed of said hybrid and line having been deposited under ATCC Accession Number PTA-120512 and ATCC Accession Number PTA-120511, respectively, and wherein the plant otherwise comprises all of the morphological and physiological characteristics of squash hybrid SV0116YL or squash line ZGN-EH10004.

21. The plant of claim 20, wherein the transgene confers a trait selected from the group consisting of male sterility, herbicide tolerance, insect resistance, pest resistance, disease resistance, modified fatty acid metabolism, environmental stress tolerance, modified carbohydrate metabolism and modified protein metabolism.

22. A plant produced by introducing a single locus conversion into a plant of squash line ZGN-EH10004, wherein the converted plant comprises the single locus conversion and otherwise comprises all of the morphological and physiological characteristics of squash line ZGN-EH10004, a sample of seed of squash line ZGN-EH10004 having been deposited under ATCC Accession Number PTA-120511.

23. The plant of claim 22, wherein the single locus conversion confers a trait selected from the group consisting of male sterility, herbicide tolerance, insect resistance, pest resistance, disease resistance, modified fatty acid metabolism, environmental stress tolerance, modified carbohydrate metabolism and modified protein metabolism.

24. A method for producing a seed of a squash plant derived from at least one of squash hybrid SV0116YL, or squash line ZGN-EH10004, said method comprising the steps of:
(a) crossing a squash plant of hybrid SV0116YL, or line ZGN-EH10004 with itself or a second squash plant; a sample of seed of said hybrid and line having been deposited under ATCC Accession Number PTA-120512, and ATCC Accession Number PTA-120511, respectively; and
(b) allowing seed of a hybrid SV0116YL, or line ZGN-EH10004-derived squash plant to form.

25. The method of claim 24, further comprising the steps of:
(c) selfing a plant grown from said hybrid SV0116YL, or ZGN-EH10004-derived squash seed to yield additional hybrid SV0116YL, or line ZGN-EH10004-derived squash seed;
(d) growing said additional hybrid SV0116YL, or line ZGN-EH10004-derived squash seed of step (c) to yield additional hybrid SV0116YL, or line ZGN-EH10004-derived squash plants; and
(e) repeating the crossing and growing steps of (c) and (d) to generate at least a first further hybrid SV0116YL, or line ZGN-EH10004-derived squash plant.

26. The method of claim 24, wherein the second squash plant is of an inbred squash line.

27. The method of claim 24, comprising crossing line ZGN-EH10004 with line LEB-EH-08-874, a sample of seed of said lines having been deposited under ATCC Accession Number PTA-120511, and ATCC Accession Number PTA-12119, respectively.

28. The method of claim 25, further comprising:
(f) crossing the further hybrid SV0116YL, or ZGN-EH10004-derived squash plant with a second squash plant to produce seed of a hybrid progeny plant.

29. A plant part of the plant of claim 7, wherein the plant part comprises a cell of squash hybrid SV0116YL.

30. The plant part of claim 29, further defined as a leaf, an ovule, pollen, a fruit, or a cell.

31. A method of producing a squash seed, said method comprising crossing the plant of claim 1 with itself or a second squash plant and allowing seed to form.

32. A method of producing a squash, said method comprising:
(a) obtaining the plant according to claim 1, wherein the plant has been cultivated to maturity; and
(b) collecting a squash from the plant.

33. A method of producing a plant of squash hybrid SV0116YL comprising a single locus conversion, the method comprising crossing a plant of line LEB-EH-08-874 with a plant of line ZGN-EH10004, wherein at least one of said lines comprises a single locus conversion, a sample of seed of said lines LEB-EH-08-874 and ZGN-EH10004 having been deposited under ATCC Accession No. PTA-12119 and ATCC Accession Number PTA-120511, respectively.