Note: Descriptions are shown in the official language in which they were submitted.
~ l~ q ~
Sunflower Products Having Lower Levels ~;
of Saturated Fatty Acids
Thomas C. Heaton
Glenn S. Cole ;~
Barry A. Martin
Background of the Invention
This invention relates to a novel sunflower (Helianthus
spp.) plant, to products obtained from the novel plant and to
methods of producing the sunflower products.
The cultivated sunflower (genus Helianthus annuus L.) is
a major worldwide source of vegetable oil. In the United
States, approximately 4 ~illion acres are planted in
sunflowers annually, primarily in the Dakotas and Minnesota.
The very rapid expansion over the last decade of acreage
planted in sunflower in the United States is due in part to
several important developments in the field of sunflower
breeding and varietal improvement. One significant dev~lop~
ment was the discovery of cytoplasmic male sterility and genes
for fertility restoration, a discovery that allowed the pro-
duction of hybrid sunflowers. ~he hybrids thus produced were
introduced during the early 1970's.
A description of cytoplasmic male sterility (CMS) and
genetic fertility restoration in sunflowers is presented by
Fick, "Breeding and Genetics", in Sunflower Science and Tech- -`~
25 noloqv Z79-338 (J.F. Carter ed. 1978).
Sunflower oil is comprised primarily of palmitic,
stearic, oleic, linoleic and linolenic acids. While other
unusual ~ ~
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fatty acids exist in plants, palmitic, stearic, oleic, lino
leic, and linolenic acids comprise about 88% of the fatt~
acids present in the world production of vegetable oils.
(Harwood, J.L. Plant Acyl Lipids: Structure, Distribution and
Analysis, 4 Lipids: Structure and Function, P.K. Stumpf and
E.E. Conn ed. (1988)). Palmitic and stearic acids are
saturated fatty acids that have been demonstrated in certain
studies to contribute to an increase in the plasma cholesterol
level, a factor in coronary heart disease. Vegetable oils
high in unsaturated fatty acids, such as oleic and linoleic
acids, may have the ability to lower plasma cholesterol
according to recent studies. Saturated fatty acids also have
higher melting points in general than unsaturated fatty acids
of the same carbon number, which contributes to cold tolerance
problems in foodstuffs and can contribute to a waxy or greasy
mouthfeel. It is also known that food products made from fats
and oils having less than 3% saturated fatty acids will
typically contain less than 0.5 gram saturated fat per serving
and as a result can be labeled as containing "zero saturated
fat" under current labelling regulations. Thus, for a number
of reasons it is desirable to produce a sunflower oil having
low levels of palmitic and stearic acids and high levels of
oleic or linoleic acids.
Prior to the present invention there were no naturally
occurring sources of low saturated fatty acid sunflower oils.
The generally accepted pathway of fatty acid biosynthesis in
plants is that palmitic and stearic acids are the products of
the beta ketoacyl-ACP synthetase system which includes three
isozymes which are referred to as KAS I, II and III.
Stearoyl-ACP acid is desaturated to oleic acid which is
esterified to phosphatidylcholine then sequentially desatu-
rated to linoleic and linolenic acids (Stymne, S and Stobart,A.K. Triacylglycerol Biosynthesis, 9 The Biochemistry of
Plants: _ Comprehensive Treatise 175-214 (1987) and Stumpf,
P.K., Biosynthesis of Fatty Acids in Higher Plants, Oil Crops
of the World 38-62 (1989), Previously published research in
.. ~-: .. .. ,. ~,
-. 2 ~
sunflower has e~phasized generally the alteration of the per-
centage of oleic or linoleic acids. The relative proporti ;
of oleîc and linoleic acids can be environmentally infiuenced
(Kinman, M.L., and F.R. Earle., "Agronomic Performance and
Chemical Composition of the Seed of Sunflower Hybrids and
Introduced Varieties," Crop Science 4:417-420 (1964~; Putt,
.
E.D., B.M. Craig, and R.B. Carson. "Variation in Composition
of Sunflower Oil from Composite Samples and Single Seeds of
Varieties and Inbred Lines, J. Am. Oil Chem. Soc. 46:126-129
(1969); Seiler, G.J., "Variation in Oil and Oil Quality of
Wild Annual Sunflower (~ _S ~ L.) Populations in a
Uniform Environment," 10th International Sunflower Conference,
March 14-18, 1982. Surfers' Paradise, Australia. p. 212-215;
Seiler, G.J., "Effect of Genotype, Flowering Date, and
Environment on Oil Content and Oil Quality of Wild Sunflower
Seed," Crop Science, 23: 1063-1068 (1983); Seiler, G.J.,
"Interrelation of Fatty Acids in Oil of Wild Annual Sunflower
(Helianthus annuus L.)" Proceedings of the XI International
Sunflower Conference._ March 10-13, 1985. Mar del Plata,
Argentina. p. 529- 534; or have been shown in certain geno-
types to be inherited in stable manner (Soldatov, R.I.,
~Chemical Mutagenesis in Sunflower Breeding, n International
Proceedinqs, 7th International Sunflower Conference,
Krasnodar, U.S.S.R., 27 June - 3 July, 1976. International
Sunflower Association Vlaardingen, p. 352-357, The
Netherlands; Karachenko, L.N., "Genotypic and Phenotypic
Mechanisms Ensuring Regulation of Fatty Acid Biosynthesis in
Sunflower Seeds," Fiziologiva Rastenii (Russian) 26: 1226-1232
(1979); Fick, G.N., "Breeding and Genetics," Sunflower Science
and Technoloqy, Carter, Jack F. (ed.). 1978. Urie, A.L.,
"Inheritance of Very High Oleic Acid Content in Sunflower,"
Proc. Sunflower Research Wockshop. Bismarck, ND. 1 February,
~.. ~,. . .
1984. National Sunflower Association. Bismarck, ND. p.9-10; ~-
Miller, J.F. and D.C. zimmerman. "Inheritance of High Oleic ,~
Fatty Acid Content in Sunflower." Proc. Sunflower Research
Workshop. Fargo, ND. 26 January, 1983. National Sunflower ; -
- 3 ~
'
~ ' '; . . ' . ' ' '
Association. Bismarck, ND. p. 10; Urie, A. L. "Inheritanc-
of High Oleic Acid in Sunflower." CroP Science 25:986-98
(1985); Simpson, B.W. and D. L. George, "Potential for
Selection of Fatty Acids on a Single Seed Basis in Sunflower
(Helianthus annuus L.)." Proceedinqs of the XI Int$rnational
Sunflower Conference, March 10-13, 1985. Mar del Plata,
Argentina. p 791-796; Miller, J.F. D.C. Zimmerman, and B.A.
Vick, "Genetic Control of High Oleic Acid Content in Sunflower
Oil," Crop Science 27:923-926 (1987); George, D.L., B.W.
Simpson, and C.M. McLeod. "Proposed Development of a High
Linoleic Acid Sunflower Hybrid." Proceedinqs of the 12th
International Sunflower Conference. July 25-29, 1988. Novi
Sad, Yugoslavia. p 448-543; Simpson B.W., C.M. McLeod and D.L.
George. "Selections for High Linoleic Acid Content in
Sunflower (Helianthus annus L.)." Aust. J. of Ex~er. Aaric.
29:233-239 (1989). Recent research has claimed that the level
of palmitic acid in sunflower oil can be increased to as high
as 40.2% of the total oil (Ivanov, P, D. Petakov, V. Nikolova,
and E. Pentchev, "Sunflower Breeding for High Palmitic Acid
Content in the Oil." Proceedinqs of the 12th International
Sunflower Conference. Vol II. July 25-29, 1988. Novi Sad,
Yugoslavia. p 463-465). The invention disclosed here pertains
to the proportion of palmitic and stearic acids relative to
the other major fatty acids in sunflower oil: oleic and
linoleic acids.
: :.,.:,.
SummarY of the Invention
This invention relates to the use of a cytoplasmically~
inherited trait that lowers sunflower seed saturated fatty ;
acid content. This trait was isolated in specific sunflower -~-
cultivars and is easily transferable to any other sunflower
line when the plant carrying the trait is used as the female
parent in a sunflower cross so that the progeny inherit the
maternal cytoplasm of the plant carrying the trait.
- ~A - .:
~ .,
The history of the isolation of this unknown and uniqu~
determinant is described in this application (see pages 6-8).
To summarize, originally, sunflower line VK9G was crossed with
pollen of variety "Pervenets" (page 6, lines 29-30). Inbred
selection was carried out for six generations and maintainer
as well as cytoplasmically male sterile counterparts were
developed. The trait was first noted in the CMS counterparts,
8904W04F and 8904W06F (Table 1). Sister lines grown in Hawaii ~ ri,,~
demonstrated the same unique results. The male sterile forms
produced seed oil with considerably reduced levels of
saturated fatty acids independent of environmental effects.
Because the invention relates to the use of a cytoplasmic
trait, its effects are inherited only when the plant carrying
this trait is used as the female parent. However, this novel '~
15 invention is not linked to male sterility in sunflowers. ;~
Table 3 shows that hybrids as well and inbreds can be produced
with low total saturated fatty acids, and when restorers are `
utilized so that the hybrids thus produced are self~
pollinating, seeds of the F2 generation produce oils with low ~ - -
saturated fatty acids when self-pollinated. Compared to the
standard (normal plants lacking the trait), both palmitic and
stearic acid levels are reduced. Thus, male sterility is a
convenient but entirely unnecessary feature of this invention.
Furthermore, Table 3 shows the same results occur across
different environments, demonstrating that there are few, if
any, environmental effects on expression of this trait. This
phenomenon is unlike any known nuclear trait affecting fatty
acid composition of seed oil, including the high-oleic ;~
varieties. ~-
The cytoplasm carrying this trait can be incorporated
into any sunflower variety or cultivar, whether fertile, male i~
sterile, or maintainer, tall or dwarf, high-yielding or low.
Any sunflower breeder skilled in the art provided the ~ ~ ;
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knowledge of a plant incorporating this trait and the presen'
disclosure, would be enabled to make and utilize the instan~
invention.
In plants, the female and male parents contribute equal
amount of chromosonal DNA to the zygote. In addition, it is ;~ -~
generally accepted that the female parent contributes the
cytoplasm, including all of the DNA-bearing cytoplasmic
organelles to the zygote. The pollen parent does not ~--
contribute to the cytoplasm of the zygote. The plant on which
10 the seed is borne is called the maternal parent or seed '''~;''`r''`''~'"'`parent. Therefore, all cells of the plant derived from seeds ~ -
of a cross have the cytoplasm of the maternal parent and equal
amounts of nuclear DNA from the female and male parents. In - -
plants where self pollination can occur, inbred lines can be
made such that plants have the cytoplasm of the maternal
parent as well as 100% of the nuclear DNA of the maternal
parent. Conversely, a breeder can start with a female
parental line having nuclear genotype Nf and cytoplasm Cf, a .
male parental line having nuclear genotype Nm and cytoplasm
20 Cm~ and using backcross techniques in which pollen from the ;~
male is used as the recurrent parent, produce a line having ~ ~
cytoplasm Cf and nuclear genotype Nm. : ~ ~.
...,..;-.:
The classical method for the determination of cytoplasmic
inheritance is to note whether there is a difference in the
trait in progeny from reciprocal crosses of two plant types.
In this case, pollen from plant A is used as the male parent
on plant B, and in the reciprocal cross, pollen from plant B
is used on plant A. The result is seed in which the cytoplasm
is from either A or B depending on which was used as the
maternal parent, and the DNA in the nucleus is identical. If
the inheritance of a trait continually follows a particular
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.
maternal line regardless of the nuclear genotype, the~
cytoplasmic inheritance is shown in that maternal parent line. ;
The present invention is the first report that a
cytoplasmic trait~s) can affect fatty acid composition in
sunflower and may be the first report of such for any oilseed.
To demonstrate this, applicant points to Tables 3, 5, and 6
which show the reduction of total saturated fatty acids in
hybrids wherein inbreds or hybrids carrying this novel
cytoplasmic trait were used as female parents and the nuclear
10 content of the progeny was either 50% or 75% distinct from -s
that of the original inbred carrying this trail. In Table 6,
particularly, sterile hybrids were made using a plant
designated 8904W06F (carrying the determinant) by crossing it
to a maintainer PHA078 (lacking the determinant), used as a
male parent. The entire cytoplasm of the hybrid, but only
one-half of the nuclear material, was inherited from 8904W06F.
The genetic content of the F1 nucleus was further diluted by
crossing this hybrid with either of three other varieties `~
(lacking the cytoplasmic trait of this invention) all of which
20 were used as the pollen donors. In other words, three new ~ ~
hybrids were made using the F1 hybrid as the female plant. ` -
Thus the nucleus of any of these F2 progeny was 75~ distinct
from 8904W06F but the cytoplasm was genetically identical. ~ ;~
~1~ of the F2 progeny produced seeds with low total saturated
25 fatty acids demonstrating the presence of a ~ -~
heritablecytoplasmic determinant. No segregation of this `
trait was observed in any of the crosses.
The action of the cytoplasmically-inherited trait of this
invention is independent of the nuclear genotype. Both normal
sunflower lines, having seed with high content of linoleic or
oleic fatty acid, have reduced saturated fatty acid content in ~ ~`p~
their seed oil when crossed into a SFPET1 cytoplasm. Since
fatty acid content is the arithmetic total of all fatty acids,
a decrease in saturated acids would result in a proportional
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increase in unsaturated fatty acids. Coincident wit~
decreased levels of saturated fatty acids in oil from seeds OL
plants of this invention, is a further increase in unsaturated : -
fatty acids.
~' ' ~ ' . . . ' ~
The cytoplasmic trait affecting saturated fatty acids in
seeds of this invention can be used in conjunction with
different nuclear genotypes that also control fatty acid
synthesis. The combination of this cytoplasmic trait with
exîsting and yet to be discovered nuclear genetic variants
offers unique fatty acid compositions in sunflower seed oil.
The present invention provides a sunflower seed that has
a total saturated fatty acid content of approximately 6% or
less.
- , ....
The present invention provides a new sunflower which is
true-breeding under a wide variety of growing conditions for
the trait of low saturated fatty acid content. This trait can
be identified and characterized by restriction fragment length
polymorphism analysis of the genome of the sunflower varieties
provided by this invention.
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This invention provides a new sunflower plant that can b~
used efficiently to produce parent lines and hybrid~
possessing desirable agronomic traits in combination with a
low content of saturated fatty acids.
This invention also provides a method for producing a
hybrid sunflower that has seed which has a total saturated
fatty acid content of approximately 10% or less.
This invention provides a novel sunflower oil that has a
total level of saturated fatty acids of 10% or less.
In accomplishing the foregoing, there has been provided,
in accordance with the present invention, a sunflower seed
having a total saturated fatty acid content of 10% or less.
There has also been provided, in accordance with another
aspect of the present invention, a hybrid sunflower which is
cytoplasmic male sterile, or alternatively, which comprises a
genetic determinant encoding fertility restoration, and which,
in addition, produces seed the triglycerides of which have a
total saturated fatty acid content of 10% or less.
In accordance with yet another aspect of the present
invention, there has been provided a sunflower oil which as
obtained fromethe sunflower seed contains approximately 10%
total saturated fatty acids or less relative to its total
fatty acid content.
Other 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 specific examples, while indicating preferred
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 area from this detailed description.
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Detailed Description --
In the description and examples that follow, a number of
terms are used herein. In order to provide a clear and
consistent understanding of the specification and claims,
including the scope to be given such terms, the following ~;~
definitions are provided~
Sunflower Seed: Botanically referred to as an "achene",
comprised of the pericarp and embryo.
Maintainer Line: A male fertile version of an inbred
10 sunflower line.
Cytoplasmic male sterile (CMS) plant or inbred line: A
sunflower line that produces no viable pollen is called male
sterile. Male sterility is inherited maternally, ie. the
male sterile plant is used as the female parent in a cross
15 with pollen from another sunflower. CMS lines are produced by
crossing a maintainer line with a sunflower plant with the
cytoplasmic male sterility trait and then backcrossing to the
maintainer line until a male sterile line that is homologous -
to the maintainer line in all other respects is developed. ;~
20 CMS lines are also referred to as female lines.
Restorer Line: A line possessing the gene or genes to
restore male fertility or viable pollen to a sunflower hybrid
or inbred line and progeny having a maternal cytoplasm that ~ ~
conditions male sterility. This term is also discussed in the h.. ,
25 literature. See, for example, the Fick article identified
above. :
Plant Breedinq
To produce the novel sunflower of the present invention
30 sunflower line VK9G was emasculated and crossed with pollen of
variety 'Pervenets.' Inbred line VK9G is a male~fertile sun~
flower line that produces seeds which have a normal fatty acid
composition. 'Pervenets' is an open-pollinated (heterogenous) `
variety introduced from the USSR in the 1970's (Soldatov,
35 1976) with high levels of oleic acid. Similar breeding ~- ~ F
methods are described in Fernandez-Martinez, J., Dominguez-
- G
Giminez, J. and Jiminez-Ramirez, A., Breeding for High Content
of Oleic Acid in Sunflower ~Helianthus annuus L.) Oil, Helia
Nr. Scientific sulletin- of the F.A.O. Research Network on -~
Sunflower 11-15 1988); Fick, G.N., Sunflower, Oil cro~s of
the World Ch. 14 pp 301-318 (1989); Knowles, P.F. Genetics and
Breeding of Oil Crops, Oil crops of the World Ch. 12 pp. 260-
282 (1989). Pedigree selection was made for six generations.
A fatty acid analysis was done on the seeds of maintainer
lines that had been inbred for six generations (~) from the
pedigree VK9GXPervenets. Also the CMS counterparts of the
maintainer lines were analyzed for fatty acid composition. At
that time, the maintainer lines had been backcrossed into the
CMS background three times so the CMS counterpart was 87.5%
homologous to the maintainer lines.
Breeding with the lines was continued in Hawaii by self
pollinating heads (capitulae) of the maintainer lines and
crossing pollen from individual heads of the maintainer lines
to their respective sterile counterparts so the identity of
each crossed pair was preserved. Fatty acid analyses were
done on seeds of the maintainer lines and selected seeds were
planted again in Woodland, CA. Crosses between the maintainer
line and the male sterile counterpart were made in pairs and
selections were made for good agronomic plant traits. Fatty
acid analysis were performed on a bulk of five seeds from each
pair of maintainer and sterile lines derived from that cross.
The maintainer selections were (VK9G/PERV)XC111211 and
(VK9G/PERV)XC111121. Those lines were the F7 generation of
inbreeding and were referred to as 8904W04G and 8904W06G. The
male sterile counterpart was greater than 97~ homologous to
its respective maintainer, having been crossed 5 times. At
that time it was noted for the first time that the seed from
the male sterile form of each maintainer had lower total
saturated fatty acids than the maintainer. The results of
this analysis are shown in Table 1.
-- 7
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able 1. ;~
Unsaturates Saturates
Entry Name Source % 18:2 % 18:1 % 16:0 % 18:0
28 8904W04F 8WB9-3 4-1 4.0 89.9 3.-9 2.2
29 8904W04G 8WB9-4 4-1 3.9 86.2 3.5 6.4
32 8904W06F 8WB9-3 6-1 3.7 90.4 - 2.0 1.9
33 8904W06G 8WB9-4 6-1 3.2 86.6 2.2 5.9
Pairs of sterile and maintainer sister lines from the
Woodland nursery were sent to Kekaha, Hawaii for winter
nursery breeding. Additional pairs of maintainer and CMS
counterparts were crossed. Seeds from those crosses were
analyzed. Once again the sterile forms of the maintainer
15 lines had lower total saturated fatty acids. Saturated fatty `
acids of the sterile lines comprised less than 7% of the total
oil content. Maintainer line selections of 8904W03G (a sister -
line selection of 8904W05G and 8904W06G) and 8904W06G had
total saturated fatty acid amounts between 8.3 and 10.2% of
the total oil content which was higher than their sterile
counterparts ~Table 2.).
The fact that total saturated fatty acids in the sterile
forms were less than that of the corresponding maintainers in
the Hawaii test as well as at the Woodland test demonstrated
that a nucleo-cytoplasmic interaction in those lines affected
fatty acid synthesis. Specifically, there was a demonstrated
cytoplasmic influence on fatty acid synthesis such that the
total stearic and palmitic acid content is less in seed of the
sterile form (F) than in the paired maintainer line (G). The
result is less total saturated fatty acids in the sterile
line. This trait is inherited from generation to generation
across different environments demonstrating that low total h `~
content of saturated fatty acids in these sunflower lines is
genetically determined in a predictable and heritable manner.
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Table 2.
Entry Name Source Unsaturates% Saturates%
18:2 18:3 18~116:0 18:0 ;-
60 8904W03F L8-91811kl4.3 0.6 88.1 4.8 2.2
61 8904W03G L8-92811kl3.0 0.6 87.9 3.5 5.0
62 8904W03F L8-91811k24.2 0.5 88.6 4.9 1.8
63 8904W03G L8-92811k22.9 0.6 88.2 3.8 4.5 ;
64 8904W03F L8-91811k34.6 0.5 88.1 4.9 2.0
65 8904W03G L8-92811k33.0 0.6 8~.4 3.7 5.3
66 8904W06F L8-91812kl0.2 0.2 88.9 4.6 2.1
67 8904W06G L8-92812kl0.4 0.1 86.1 3.8 6.9
68 890~W06F L8-91812k34.0 1.4 88.1 4.4 2.1
69 8904W06G L8-92812k33.2 0.3 80.7 5.1 5.1 ~ *
In the Woodland nursery, the sterile counterparts of
8904W03G and 8904W06G were crossed with restorer lines. That -
hybrid seed was distributed to sunflower research stations for
testing in California, France, and Argentina. Hybrids were
grown, and bags were placed on heads prior to flowering to
force self pollination of the hybrid and prevent contamina~
tion. Seeds from individual heads of those hybrids were
analyzed for fatty acid composition. It was found that oil in ;
bulked seeds of individual heads from hybrids made with the
female (male sterile) inbreds 89W04W03F and 89W04W06F was low
,J~ ' .
in total saturated fatty acids (Table 3.). The fact that
individual heads had seeds whose oil in a bulk sample had low
levels of saturated fatty acid demonstrated that such levels
can be produced in a hybrid as well as parental inbred lines.
Moreover, the low total saturates were obtained in three
different sunflower growing environments. It was demonstrated
that these male sterile lines when used as female parents
2 1 2 ~ 6 2 ~
produced low saturated fatty acids in hybrids grown in typical
sunflower crop production areas.
Table 3.
Result~ of fatty acid analyses of co~posite seed samples from
individual self pollinated heads from hybrids ~ade using
8904W06F and 8904w03F ac the female parent grown in three
environments.
Location Hybrid Head Sterile Unsaturate~% Saturates%
18:218:116:0 18:0 ~ ..
Montech,
France
8W1070 Bulk 8904W06F 2.194.1 2.8 1.0
Woodland,
15 California :~
8W1070 1 8904W06F4.4 87.6 S .4 2.3
2 4.2 89.5 S.3 1.1
3 4.5 91.1 3.5 0.9 -
4 7.0 88.2 3.6 1.2
3.8 90.8 4.5 0.9 ``
8W1075 1 8904W03F3.5 91.6 3.9 1.0
2 8.6 86.4 3.7 1.3
3 4.0 90.6 4.4 0.9
4 4.0 91.0 4.2 0.9 `~
25 Venado Tuerto,
Argentina
8W1070 1 8904W06F10.4 80.04.1 5.1 ~ ` i
2 1.7 91.43.8 3.1
3 1.6 93.53.3 1.5
4 1.9 93.63.7 0.7
8W1075 1 8904W03F1.9 92.83.6 1.7 -~
2 1.7 94.23.2 0.9
3 2.2 93.43.4 1.0
4 2.3 93.53.4 1.0 ; ';
6440
Standard Comparison 66.220.8 7.0 5.8 ~ ~
~- ' '',' ~ '
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Data from other fatty acid analyses of sunflower seed were
reviewed. A number of restorer lines were found that had
total saturated fatty acid levels of less than 5% (Table 4).
Table 4 -~
Summary of restorer lines found in fatty acid analysis screen~
ing that have less than 5% total saturated fatty acid levels.
LineSelection %16:0 ~18:0Total Sat %18:1 %18:2
9A4W005M 1 3.0 1.6 4.6 90.8 4.5
9A4W005M 6 3.2 1.4 4.6 91.4 4.0
9A4W005M 7 3.2 1.6 4.8 94.9 2.0
9A4W005M11 3.1 1.5 4.6 91.7 3.8
9B3W006M 1 2.8 1.8 4.6 92.7 2.7
9~3W006M 3 2.9 2.0 4.9 93.2 1.8
9B3W006M11 2.6 l.0 3.6 94.4 1.9
9B3W006M12 3.2 1.5 4.7 92.4 2.8
9E-RUN737 2 2.3 1.8 4.1 95.0 1.0
- ' ' "' S`~,,J,~'
In the Hawaiian winter nursery hybrids were synthesized by
crossing female sunflower lines having seeds with less than 6%
total saturated fatty acids with male (restorer) lines having
seed with less than 6% total saturated fatty acids. Seeds ;
from the plants resulting from those crosses were planted at
Woodland, CA and Moorhead, MN. At flowering, heads of those
hybrids were bagged to assure self pollination. Heads were
harvested and individual seeds from separate identity pre- :
, ~
: .. .....
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served heads were analyzed for fatty acid composition. T~
results are presented in Table 5.
Table 5
~ ummary of total saturated fatty acid (SFA) levels in seeds of
indivi~ual ~elf pollinated heads of hybrids grown in Woodl~nd,
CA and Noorhea~, MN.
Lowest SFA~ Observation
Heads ~eeds SFA~
~ybridLocation no. no. mean sinale seeds~nale head
9W1204Moorhead 5 25 4.8 3.9 4.1
Woodland 5 25 5.2 4.0 4.3 ~ ~:
9W1224Moorhead 5 25 4.5 4.0 4.2
Woodland 5 25 5.3 4.5 4.8
9W1234Moorhead 5 25 4.6 3.8 4.2
Woodland 5 23 5.6 4.8 5.2
9W1244Moorhead 3 15 4.4 4.0 4.1 ~ ` -
Woodland 5 25 5.4 4.3 5.1 i
9W1284Moorhead 5 25 4.7 4.4 4.6
Woodland 5 24 5.9 3.9 4.8
6440Moorhead 5 25 10.4 9.0 9.8
Standard Woodland 5 25 10.5 8.8 10.5
The data in Table 5 demonstrate that inbred sunflower
lines having less than 6% total saturated fatty acids combined ~-
25 to produce hybrids whose seed had less that 6% total saturated -
fatty acids in both the Woodland and Moorhead environments.
Individual heads were produced in both environments that had
less than 5% total saturated fatty acids. In both
environments individual seeds were also produced that had less -
than 4~ total saturated fatty acids.
In order to validate the maternaL cytoplasmic effect on
the level of saturated fatty acids in this discovery, female
line 8904W06F with cytoplasm conferring low saturated fatty
acid totals was crossed with another sunflower maintainer line
PHA078. PHA078 is a proprietary inbred line having high oleic ~ --
acid content. As expected, the progeny from the cross of
- 12 ~
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8904W06F/PHA078 were sterile hybrids since PHA078 is a main-
tainer line and lacks genes for male fertility restoration c
8904Wo6F. These male sterile plants were crossed with pollen
from three other lines: PHA076, 9B3W006M, and PHA061. PHA076
and 9B3W006M are inbred lines having high oleic acid content. -~
PHA061 is an inbred line with normal fatty acid composition.
As a result of this cross, each seed from plants of
8904W06F/PHA078 could be classified as having the cytoplasm of
8904W06F and a nuclear genotype which is contributed 1/4 by
8904W06F, 1/4 PHA078, and 1/2 PHA076, 9B3W006M, or PHA061.
These seeds were analyzed for fatty acid composition. The
data are presented in Table 6. ~;
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Table 6
8ummary of total saturated fatty acid levels tSFA%) in seeds
with cytoplasmic background conferring low saturated fatty
acid traits with nuclear parentage 75~ distinct from 8904W06F.
Seeds ~F~% %Fatty A¢ids Composition
20Cross no. mean 16:0 18:0 18:1 18:2
8904W06F*PHA078/PHA07610 3.80 2.91 0.89 93.78 2.77
Lowest SFA of cross 1 3.30 2.70 0.60 94.40 2.30
8904W06F*PHA078/9B3W006M 10 3.97 3.10 0.87 94.09 1.91
Lowest SFA of cross 1 3.40 2.70 0.70 95.30 1.40
8904W06F*PHA078/PHA06110 4.03 3.07 0.96 91.46 4.47
Lowest SFA of cross 1 3.40 2.50 0.90 93.90 2.70
.
The data from Table 6 demonstrate that even when the
nuclear component of 8904W06F is diluted to 75% by four other
lines (PHA078, PHA076, 9B3W006M, and PHA061) having either
normal oil composition or higher oleic fatty acid composition,
the cytoplasmic effect caused by the maternal parent 8904W06F
is maternally transmitted and the progeny have seeds with
lower total saturated fatty acid content.
- 13 ~
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Thus, while not intending to be limited by theory, it .
~ appears that the cytoplasm contains the factors which control
for reduced saturated fatty acid levels. These cytoplasmic
factors can be transferred to offspring when the plants .
according to this invention are used as the female parent in a
cross with other sunflower cultivars, providing a method of ;~
reducing the content of saturated fatty acid moieties in any .
sunflower cultivar by conventional crossing and backcrossing.
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Deposits ~ ~-
Applicants have made available to the public without
restriction a deposit of at least 2500 seeds of a sunflower
designated 890403F with the American Type Culture Collection ~ ~r~
(ATCC), Rockville, MD 20852, W.S.A., ATCC Deposit No.75180 .
The seeds deposited with the ATCC are taken from the same
deposit maintained by Pioneer Hi-Bred International, Inc., 700
Capital Square, 400 Locust Street, Des Moines, Iowa, 50309,
since prior to the filing date of this application. The
deposit will be maintained without restrictions, at the ATCC
Depository, which is a public depository, for a period of 30
years, or five years after the most recent request, or for the
effective life of the patent, whichever is longer, and will be
replaced if it becomes nonviable during that period.
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