U.S. patent application number 13/958858 was filed with the patent office on 2013-11-28 for methods for producing a hybrid seed product.
This patent application is currently assigned to DAIRYLAND SEED CO. INC.. The applicant listed for this patent is DAIRYLAND SEED CO. INC.. Invention is credited to Paul Sun.
Application Number | 20130318649 13/958858 |
Document ID | / |
Family ID | 37028656 |
Filed Date | 2013-11-28 |
United States Patent
Application |
20130318649 |
Kind Code |
A1 |
Sun; Paul |
November 28, 2013 |
METHODS FOR PRODUCING A HYBRID SEED PRODUCT
Abstract
A method for increasing production of hybrid seed of
bee-pollinated crops, such as alfalfa and soybean at predetermined
hybridity levels. Hybrid seed is produced using female and
pollenizer plants at a selected ratio of female plants to
pollenizer plants. The female plants and the pollenizer plants are
intermingled in the hybrid seed production field. Prediction of
percentage of hybridity at various female to pollenizer ratios
allows for selection of a ratio of female plants to pollenizer
plants to provide seed at a test percentage of hybridity. The
percentage of hybridity may be increased post-harvest by employing
techniques using seed properties such as size differential, color
or density to remove a higher percentage of non-hybrid seed. The
hybrid seed product is maximized at various hybridity levels.
Planting according to subrows allows for separate harvesting of
intermingled crops. Testing the hybrid seed product provides
verification of percentage of hybridity.
Inventors: |
Sun; Paul; (Roscoe,
IL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
DAIRYLAND SEED CO. INC. |
WEST BEND |
WI |
US |
|
|
Assignee: |
DAIRYLAND SEED CO. INC.
West Bend
WI
|
Family ID: |
37028656 |
Appl. No.: |
13/958858 |
Filed: |
August 5, 2013 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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12409010 |
Mar 23, 2009 |
8502019 |
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13958858 |
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11146365 |
Jun 6, 2005 |
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12409010 |
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Current U.S.
Class: |
800/260 ;
435/6.1; 702/19 |
Current CPC
Class: |
A01H 1/02 20130101; A01H
5/10 20130101; G16B 20/00 20190201 |
Class at
Publication: |
800/260 ;
435/6.1; 702/19 |
International
Class: |
A01H 5/10 20060101
A01H005/10; G06F 19/18 20060101 G06F019/18 |
Claims
1. A method for producing a hybrid seed product of a bee-pollinated
crop, from a cross between female plants from at least one female
line and pollenizer plants from at least one pollenizer line, the
hybrid seed product having a percentage of hybridity above a
predetermined percentage of hybridity, the method comprising: (a)
selecting the at least one female line and the at least one
pollenizer line to have a seed coat color differential between seed
of the female plants and seed of the pollenizer plants; (b)
planting the at least one female line and the at least one
pollenizer line in a production ratio in a production field such
that the at least one female line and the at least one pollenizer
lines are crossed to obtain the total seed product comprising
hybrid seed and non-hybrid seed, wherein the hybrid seed and
non-hybrid seed in the production seed each have a seed coat color
distribution and the seed coat color distributions overlap by less
than twenty-five percent of the production seed; and (c) removing
at least a portion of the non-hybrid seed from the hybrid seed to
achieve a post-production increase of percentage of hybridity and
obtain the hybrid seed product having a percentage of hybridity
above the predetermined percentage of hybridity.
2. The method of claim 1 wherein the bee-pollinated crop is
soybean.
3. The method of claim 1 wherein the bee-pollinated crop is
alfalfa.
4. The method of claim 1 wherein the predetermined percentage of
hybridity comprises seventy-five percent.
5. The method of claim 1 wherein the production ratio of step (b)
is determined by selecting a ratio of the female line to the
pollenizer line to maximize yield of the total seed product in the
production field or yield of the hybrid seed product in the
production field, while maintaining the percentage of hybridity of
the hybrid seed product above the predetermined percentage of
hybridity.
6. The method of claim 1 wherein selecting the at least one female
line and the at least one pollenizer line in step (a) comprises:
selecting the at least one female line and the at least one
pollenizer line to provide a cross maintaining at least two-thirds
of female seed yield when increasing ratio of female plants to
pollenizer plants from 1:1 to 3:1 under substantially similar
environmental conditions.
7. The method of claim 1 wherein selecting the at least one female
line and the at least one pollenizer line comprises selecting a
female line having an average Pollen Production Index (P.P.I.)
between 0.0% to 0.42%.
8. The method of claim 1 wherein selecting the at least one female
line and the at least one pollenizer line comprises selecting a
pollenizer line having high self-incompatibility.
9. The method of claim 1 further comprising crossing the female
plants with the pollenizer plants at a test ratio of female plants
to pollenizer plants in a test plot to obtain a test seed product
having a test percentage of hybridity, wherein the production ratio
of female plants to pollenizer plants needed to produce the hybrid
seed product having a percentage of hybridity above seventy-five
percent is estimated from the test percentage of hybridity.
10. The method of claim 9 wherein the test percentage of hybridity
is determined by harvesting the test seed product; and determining
the proportion of hybrid seed in the test seed product.
11. The method of claim 10 wherein crossing the female plants with
the pollenizer plants at the test ratio in the test plot further
comprises planting female plants and pollenizer plants in the test
plot, the test plot having multiple rows, with the female plants
planted in a female subrow offset from the centerline of each row
of the multiple rows on a first side, and the pollenizer plants
planted in a pollenizer subrow offset from the centerline of each
row on an opposite side.
12. The method of claim 1 wherein substantially all the non-hybrid
seed is removed in step (c).
13. The method of claim 1 further comprising crossing the female
plants and the pollenizer plants at multiple test ratios of female
plants to pollenizer plants to determine a respective test
percentage of hybridity for each of the multiple test ratios.
14. The method of claim 13 wherein the production ratio of step (b)
is determined by: performing a regression analysis on the multiple
test ratios test percentage and the respective test percentage of
hybridity; and using the regression analysis to determine the
production ratio of female plants to pollenizer plants to produce
the hybrid seed product having a percentage of hybridity above
seventy-five percent.
15. The method of claim 1 further comprising verifying the
percentage of hybridity of the hybrid seed product by progeny
testing.
16. The method of claim 15 wherein verifying the percentage of
hybridity of the hybrid seed product by progeny testing further
comprises measuring the Pollen Production Index (P.P.I.) of the
production seed (P.P.I. (Production seed)), measuring the P.P.I. of
the test plot hybrid seed (P.P.I. (Test plot hybrid seed)), and
determining the percentage of hybridity of the hybrid seed product
(% hybridity) as: % Hybridity = 1 - P . P . L ( Production seed ) 1
- P . P . L ( Test plot hybrid seed ) .times. 100 %
##EQU00005##
17. The method of claim 15 further comprising using molecular
markers to identify the hybrid seed and the non-hybrid seed to
determine percentage of hybridity of the hybrid seed product.
18. The method of claim 15 wherein verifying percentage of
hybridity for the hybrid seed product further comprises: collecting
female seed data on traits of the female seed; collecting
pollenizer seed data on traits of the pollenizer seed; collecting
hybrid seed product data on traits of the hybrid seed product; and
analyzing the female seed data, the pollenizer seed data and the
hybrid seed product data to estimate a percentage of hybridity for
the hybrid seed product.
19. The method of claim 1 further comprising killing the pollenizer
plants prior to harvest to increase the percentage of hybridity of
the production seed.
20. The method of claim 1, wherein the portion of non-hybrid seed
is removed from the non-hybrid seed by a color sorter.
21. The method of claim 1, wherein the hybrid seed product is
produced at commercial levels of production.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. application Ser.
No. 12/409,010, filed Mar. 23, 2009, U.S. Pat. No. 8,502,019, which
is a continuation of U.S. application Ser. No. 11/146,365, filed
Jun. 6, 2005, abandoned, which are incorporated herein by reference
in their entireties.
BACKGROUND OF THE INVENTION
[0002] The present invention generally relates to methods for
production of a hybrid seed product, and more particularly to
methods for producing a hybrid seed product while increasing
production of hybrid seed of bee-pollinated crops at predetermined
hybridity levels, including post production adjustment of
percentage of hybridity and verification of percentage of hybridity
of the hybrid seed product after harvest.
[0003] Hybrid plant varieties offer many desirable agronomic
traits. Hybrid plant varieties often provide higher yield than
non-hybrid strains. Hybrid plant varieties may also possess higher
stress tolerance than non-hybrid varieties, enabling them to
survive in less favorable environmental conditions. Furthermore,
hybrid plant varieties provide greater efficiency in breeding
improvements. Hybridization allows for combination of desirable
agronomic traits from different strains.
[0004] While hybridization provides these desirable agronomic
traits, certain plant species present particular challenges to
hybridization. Species such as corn are easily hybridized because
the male and female reproductive organs have physical separation.
Other species have reproductive parts that are less accessible and
have little physical separation between male and female parts.
These species are more difficult to hybridize. Bee-pollinated
crops, such as alfalfa and soybeans, are examples of species that
have male and female reproductive parts that have little physical
separation because of the relatively small size of the flower.
[0005] Hybrid seed production in these species often employs
cytoplasmic male sterile plants also called female plants. Seed
that is produced by female plants from pollinations by pollenizer
plants, also called male plants, is mostly hybrid seed. Seed
produced from selfing or sibbing by the pollenizer plants is mostly
non-hybrid seed. During production of hybrid seed, employing a
higher proportion of female plants to pollenizer plants in a hybrid
seed production field increases the proportion of hybrid seed to
non-hybrid seed. However, current methods for producing hybrid seed
result in a significant decrease of seed yield for each individual
plant, when increasing the proportion of female plants to
pollenizer plants. There is therefore a need for methods for
increasing the proportion of hybrid seed to non-hybrid seed while
maintaining high overall seed yield for a hybrid seed product.
[0006] Production of hybrid varieties is subject to federal law
requirements that make a high percentage of hybridity very
desirable. 7 CFR .sctn.201.26 requires that a variety have at least
seventy-five percent hybridity to be classified as a hybrid
variety. Certain plant species, particularly bee-pollinated
species, such as alfalfa and soybeans, present challenges to
breeding plant varieties that meet this federally mandated
hybridity level. U.S. Pat. No. 3,570,181, herein incorporated by
reference, discloses a method for producing hybrid alfalfa using
cytoplasmic male sterile alfalfa plants. However, production
according to this method results in large reduction of seed yield
that makes production of hybrid seed of bee-pollinated crops
commercially impractical. U.S. Pat. No. 4,045,912, herein
incorporated by reference, discloses a method for producing seed
but does not provide for production of seed meeting federal
requirements for a hybrid variety. U.S. Pat. No. 4,045,912 is
further not concerned with verification of percentage of hybridity
at commercial levels of production. Thus, there is a need for
methods for production of hybrid seed that provide for high seed
yield while maintaining hybridity levels meeting federal
requirements.
[0007] Production of a certified hybrid alfalfa product requires
determination of percentage of hybridity. Determination of
percentage of hybridity is often accomplished using methods
employing morphological or molecular markers. However, using
molecular markers is expensive, takes significant time and is often
commercially impractical. In some species, morphological markers
require a homozygous recessive gene on one side, and dominance gene
on the other side, and are therefore difficult to employ.
Furthermore, some plant species present additional difficulties to
employing such methods. For example, tetraploid species such as
alfalfa have greater complexity in their genetics and inheritance.
Another difficulty is the small seed size of some species, such as
alfalfa. There is therefore a need for methods for determining or
verifying the percentage of hybridity of a particular hybrid that
avoid these difficulties.
[0008] Separate harvesting of female seed and pollenizer seed is
useful for determining percentage of hybridity. Prior methods of
separate harvesting for species such as alfalfa require planting
female seed and pollenizer seed in separate rows for separate
harvesting. However, production of hybrid seed for bee-pollinated
crops, such as alfalfa and soybeans, benefits from intermingling to
allow a higher percentage of cross-pollination to occur. Planting
in separate rows deceases production of hybrid seed, as
cross-pollination becomes less frequent due to the distance of the
female plants from the pollenizer plants. Where multiple female
rows are planted for every male row, such as shown in U.S. Pat. No.
3,570,181, cross-pollination becomes even less frequent. There is
therefore a need for methods for separately harvesting female seed
and pollenizer seed that allow for intermingling of female and
pollenizer plants.
[0009] Production of a hybrid seed product is furthermore a long,
expensive and labor-intensive process. Prior methods of seed
production do not provide for prediction of percentage of
hybridity. A method for predicting hybridity levels of seed
production of bee-pollinated crops would facilitate production of a
hybrid seed product meeting federal standards. This would allow for
faster, cost-effective and less labor-intensive production. There
is therefore a need for methods for predicting hybridity levels of
bee-pollinated crops.
[0010] Prior methods of seed production also do not include methods
for post-production adjustment of hybridity levels of
bee-pollinated crops. A method for adjusting hybridity levels of
bee-pollinated crops would also aid meeting hybridity standards for
production of hybrid plant varieties. Post-production adjustment of
hybridity levels also would allow for production of hybrid plant
varieties while employing breeding methods that would not normally
result in hybridity levels required by statute. There is therefore
a need for post-production methods for adjusting hybridity levels
of bee-pollinated crops.
[0011] Although hybrid plant varieties are desirable for agronomic
reasons, commercial production of many hybrid plant varieties has
not been commercially viable, particularly for bee-pollinated
crops. The present invention solves these needs and other problems
in the field of hybrid seed production by providing, in most
preferred aspects, methods for producing a hybrid seed product that
includes increasing production of hybrid seed of bee-pollinated
crops at predetermined hybridity levels, prediction of percentage
of hybridity, determination of percentage of hybridity,
post-production adjustment of percentage of hybridity, and
verification of percentage of hybridity after harvest.
SUMMARY OF THE INVENTION
[0012] The invention therefore provides a method for producing a
hybrid seed product of bee-pollinated crops, such as alfalfa and
soybean, at predetermined hybridity levels. Hybrid seed is produced
using female and pollenizer plants at a predetermined ratio of
female plants to pollenizer plants mixed together in a hybrid seed
production field. Prediction of percentage of hybridity at various
female to pollenizer ratios allows for selection of a ratio of
female plants to pollenizer plants to provide seed at a selected
percentage of hybridity. The percentage of hybridity of the hybrid
seed product may be increased after harvesting by employing
techniques using seed properties such as size differential, color
or density to remove a higher percentage of non-hybrid seed.
[0013] In other aspects of the present invention, the method
provides for maximization of seed yield at various hybridity
levels.
[0014] In further aspects of the present invention, the method
provides for separate harvesting of intermingled crops.
[0015] In further aspects of the present invention, the method
provides for verification of percentage of hybridity of a hybrid
product.
[0016] These and further objects and advantages of the present
invention will become clearer in light of the following detailed
description of an illustrative embodiment of this invention.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] The illustrative embodiment may best be described by
reference to the accompanying drawings where:
[0018] FIG. 1 shows a flow diagram of one embodiment of the methods
for producing a hybrid seed product.
[0019] FIG. 2a shows a regression curve showing percentage
hybridity versus female to pollenizer ratio for a single cross.
[0020] FIG. 2b shows a regression curve showing percentage
hybridity versus female to pollenizer ratio for multiple
crosses.
[0021] FIG. 3 shows a plot planted according to the subrow method
of the invention.
[0022] All figures are drawn for ease of explanation of the basic
teachings of the present invention only; the extensions of the
figures with respect to number, position, relationship, and
dimensions of the parts to form the preferred embodiment will be
explained or will be within the skill of the art after the
following description has been read and understood. Further, the
exact measurements and measurement proportions to conform to
specific percentages, sizes, and similar requirements will likewise
be within the skill of the art after the following description has
been read and understood. Values provided are representative and
are utilized to facilitate the description of the preferred
embodiment.
[0023] Where used in the various figures of the drawings, the same
numerals designate the same or similar parts. Furthermore, when the
terms "upper," "lower," "side," "end," "bottom," "first," "second,"
"laterally," "longitudinally," "row," "column," "array," and
similar terms are used herein, it should be understood that these
terms have reference only to the structure shown in the drawings as
it would appear to a person viewing the drawings and are utilized
only to facilitate describing the illustrative embodiment.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
[0024] The present invention provides methods for producing a
hybrid seed product of bee-pollinated crops such as alfalfa and
soybean. FIG. 1 shows a flow diagram of one example embodiment of
these methods. The method includes selection of a female line and a
pollenizer line 10. These lines are planted in test plots in
selected ratios of female plants to pollenizer plants 20. Data is
collected from the test plots 30. Data analysis provides
information on characteristics such as: seed size distribution for
each line and seed yield of female and pollenizer plants in
response to the selected ratios of female plants to pollenizer
plants 40. In the preferred embodiments, the data analysis can
provide for further selection of a female line or lines and further
selection of a pollenizer line or lines for production of the
hybrid seed product. In one example embodiment, the data analysis
40 may indicate a combination of a female line and pollenizer line
having a high seed yield.
[0025] In this example embodiment, analysis of seed size
distribution determines a procedure for producing the hybrid seed
product. If the seed size distribution of the female plants and the
pollenizer plants have a small amount of overlap 50, then a female
to pollenizer ratio is selected using the data analysis to produce
the maximum quantity of seed 60. In one example embodiment,
selection of a female to pollenizer ratio to produce the maximum
quantity of seed occurs when the overlap in seed size distribution
involves less than 25% of the seed product of the cross between the
selected female line and the selected pollenizer line. Subsequent
seed size screening removes enough non-hybrid seed to achieve a
targeted percentage of hybridity in the hybrid seed product. Where
there is little or no overlap in seed size distribution,
substantially all of the non-hybrid seed may be removed 70.
[0026] If the seed size distribution of the female plants and seed
size distribution of the pollenizer plants have significant
overlap, but are not substantially the same 80, then statistical
analysis on the respective seed size distributions determines the
percentage of hybridity that can be gained by screening out seed
within a selected size range 90. Selection of a female to
pollenizer ratio for a production field takes into account both the
hybridity level of the selected female to pollenizer ratios in the
test plots, and the gain in hybridity level attained by screening
out seed in a selected size range 100. In the preferred embodiment,
the ratio producing the maximum quantity of seed after post-harvest
screening to achieve a targeted hybridity level is selected for use
in the production field.
[0027] If the seed size distribution of the female plants and the
seed size distribution of the pollenizer plants are substantially
the same 110, then a female to pollenizer ratio is selected for the
production field using data from the test plots that will achieve
the targeted hybridity level 120.
[0028] The selected female line and the selected pollenizer line
are then planted in the production field in the selected female to
pollenizer ratio to produce the hybrid seed product 130. After
production, the percentage of hybridity of the hybrid seed product
is verified 140.
[0029] With respect to the selection of lines 10 for the hybrid
seed product, in the preferred embodiment of the methods of the
invention, the production of a hybrid seed product includes
selection of one or more female lines, also referred to as an "A
line." A female line is a cytoplasmic male sterile line, a genic
male sterile line, or an induced male sterile line by chemicals or
biotechnology. These lines have a condition in which pollen is
absent or non-functional in flowering plants. The methods also
include the selection of one or more pollenizer lines, also
referred to as a "male line" or "C line." The pollenizer line is a
male fertile line having a condition in which pollen is produced
and functional in flowering plants, with the female part either
functional or not functional.
[0030] In the preferred embodiment, selection of the female line
and the pollenizer line takes into account both agronomic factors
and factors that affect percentage of hybridity and seed yield. In
an example embodiment, these factors include female to pollenizer
ratio, seed yield from pollenizer plants, seed yield from female
plants, female to pollenizer seed yield differential, female to
pollenizer seed size differential, high seed yield with increasing
female to pollenizer plant ratio, low P.P.I. (Pollen Production
Index), large seed size, pollination power in pollenizer plants and
high self-compatibility in pollenizer plants. In further aspects of
the invention, these factors include desirable agronomic traits
such as disease resistance, insect resistance and forage yield.
[0031] The selected lines can then be planted in test plots 20 for
data collection 30 and data analysis 40. In the preferred
embodiment, data on seed yielding ability, pollination power,
pollen production index (P.P.I.), seed size distribution for each
line and seed yield of female and pollenizer plants in response to
the selected ratios of female plants to pollenizer plants is
collected along with other characteristics.
[0032] The data collection 30 and data analysis 40 of the present
invention demonstrate that individual female lines can have
different seed yielding abilities. Table 1 illustrates test results
of seed yield for four female lines with different P.P.I., with
each female line crossed with five different pollenizers.
TABLE-US-00001 TABLE 1 Seed Yield of Female Plants with Different
P.P.I. Female Line Average seed Yield (AXB) P.P.I. (Total wt. of 4
rows in grams) Female1 0.013 328.6 Female2 0.025 402.1 Female3
0.046 318.2 Female4 0.085 369.6
[0033] Table 1 illustrates the results of testing under the
following conditions: [0034] Selection of four female lines with
different P.P.I. and five pollenizer lines with different
pollinating abilities for diallel mating in three female to
pollenizer ratios, 70:30, 75:25 and 80:20 at two locations,
Sloughouse, Calif., and Homedale, Id. p1 Split plot designs with
two replications were used at the two locations. Each location
included 120 plots, with each plot having six rows and being 20
feet long. The center four rows included a mixture of both female
and pollenizer seed in one of the tested ratios. The outside rows
included only female seed and provided for P.P.I., readings and
progeny testing.
[0035] These four female lines demonstrate different seed yielding
capabilities. As further shown in Table 1, experimentation reveals
no significant correlation between P.P.I. and seed yields for the
tested female lines in the tested female to pollenizer ratios
within the tested low P.P.I. range. In this experiment, each female
line was crossed with five male lines in three female to pollenizer
ratios. Thus, in a preferred embodiment of the invention, the
methods of the invention for hybrid seed production includes
selection of at least one female line having a high seed yield,
such as the line designated Female2 in the study above, without
regard for the P.P.I. of the female line. In other aspects of the
present invention, the invention may employ multiple female lines
having varying seed yield. Those skilled in the art will recognize
that selection of a female line may be done according to various
agronomic factors within the spirit and scope of the invention.
[0036] In the preferred embodiment of the present invention,
selection of a pollenizer, or male, line provides for high
pollination power. Thus, a measure of pollination power provides a
measure of average female line seed yield from pollination of the
selected pollenizer line. Tables 2 and 3 show representative
measures of pollination power for five male lines.
TABLE-US-00002 TABLE 2 Seed Yield of Pollenizers Pollenizer Seed
Yield (grams/plant) Male1 361.5 Male2 333.5 Male3 360.8 Male4 440.7
Male5 276.6
[0037] Table 2 shows data collected according to the teachings of
the present invention on average seed yield for five pollenizers
crossed with four different females in three female to pollenizer
ratios: 80:20, 75:25 and 70:30. Seed were harvested in bulk,
comprising a mixture of both hybrid and pollenizer seed. Seed
yields axe the mean of twenty-four plots. The Male5 pollenizer line
had the white flower trait.
[0038] Testing according to the teachings of the present invention
indicates that different pollenizers have different seed yielding
abilities and varying pollinating power with respect to female
plant seed yield. In the study performed above, the Male4
pollenizer caused the female plants to produce the most seed, and
the Male5 pollenizer caused the female plants to produce the least
seed. These results are statistically significant at the 1%
level.
TABLE-US-00003 TABLE 3 Average Seed Yield of Each Female with
Specific Pollenizer (Male) at Three Female to Male Ratios (70:30,
75:25, and 80:20) Male Male4 Male5 Female Male1 Male2 Male3 (High
seed) (Low seed) Female1 316.8 339.2 316.8 383.8 286.7 Female2
431.2 392.5 361.5 534.3 290.8 (high seed yielder) Female3 297.0
298.5 358.0 402.3 235.3 (low seed yielder) Female4 401.2 303.7
406.8 442.8 293.5
[0039] Testing according to the teachings of the present invention
confirm that crossing females having high seed yield with
pollenizers having high seed yield results in a cross with the
highest seed yield. Likewise, crossing females having low seed
yield with pollenizers having low seed yield results in a cross
with the lowest seed yield. Table 3 summarizes the results of this
testing.
[0040] According to the further teachings of the present invention,
the percentage of hybridity of the hybrid seed product will be
highest when using a male line with high pollination power but low
seed yield. This provides for a higher percentage in the production
field of hybrid seed by lowering production of non-hybrid seed. In
the preferred form of the invention, the method includes selection
of a male line with low seed yield but high pollination power for
crossing with a female line with high seed yield in hybrid seed
production. In another aspect of the preferred embodiment of the
invention, pollenizers having high self-incompatibility contribute
to a hybrid seed product having less selfed seed than outcross
seed. As with selection of the female line, in other aspects of the
present invention, the method of the invention may employ multiple
pollenizer lines having varying pollination power. Those skilled in
the art will recognize that selection of a pollenizer line having
lower pollination power and lower self-incompatibility is within
the spirit and scope of the invention, and that selection of a
pollenizer line may be done according to various agronomic
factors.
TABLE-US-00004 TABLE 4 Seed Yield Decrease with Increase Female and
Decrease Pollenizer (Male) in Female to Male Ratios
Female:Pollenizer(male) 70:30 75:25 80:20 Seed Yield 375.2 370.1
318.6 (Hybrid seed and pollenizer seed) % reduction from 70:30 1.4
15.1 seed production level
[0041] In the most preferred embodiment of the invention, the
method for production of hybrid seed further incorporates selection
of female to pollenizer ratios. Testing according to the teachings
of the present invention demonstrates that female to pollenizer
ratio affects seed yield, as shown in Table 4. This testing further
indicates that the effect of female to pollenizer ratio is not
linear. In one example embodiment, experimentation on selected
female and pollenizer lines demonstrates an average 1.4% drop in
seed yield between a 70:30 and a 75:25 female to pollenizer ratio.
However, testing demonstrated an average 15.1% drop in seed yield
between a 75:25 and an 80:20 female to pollenizer ratio. Therefore,
for the tested lines, much greater gains in seed yield occur when
going from an 80:20 female to pollenizer ratio to a 75:25 ratio
than from a 75:25 ratio to a 70:30 ratio. Table 4 summarizes the
average decreases in seed yield with increasing female to
pollenizer ratios for the tested varieties. Furthermore, testing
according to the teachings of the present invention further
demonstrates that the decrease in seed yield will vary with
different female to pollenizer crosses, as shown in Tables 6,7,8,
and 9.
[0042] According to the teachings of the present invention,
individual crosses have varying responses to female to pollenizer
ratio with respect to hybrid seed production levels and percentages
of hybridity. Thus, in the preferred form of the invention,
production of a hybrid seed product includes determining, for each
cross, the hybrid seed production level and percentage of hybridity
for a range of female to pollenizer ratios. In this preferred
embodiment, hybrid seed production employs selection of a specific
female to pollenizer ratio for maximum seed production at a desired
hybridity level for each cross of two or more lines. However, those
skilled in the art will recognize that selection of a female to
pollenizer ratio can be made for a selected cross without
determining the hybrid seed production level and percentage of
hybridity for a range of female to pollenizer ratios. For example,
in one alternate embodiment, a typical response to female to
pollenizer ratio may be used for production purposes without
determining individual response to female to pollenizer ratio. In
another alternate embodiment, where non-hybrid seed are removed to
produce the hybrid seed product, a female to pollenizer ratio
having high hybrid seed production levels, such as 70:30 or 60:40
is selected for production.
TABLE-US-00005 TABLE 5 Seed Yield of a Female Line Pollinated with
High and Low Seed Yield Pollenizers at Three Different Female to
Pollenizer Ratio's Female:Male ratio 1:1 2:1 3:1 total C1/C2 ratio
A1 .times. C1/A1 .times. C2 ratio F(A) (wt/plant in grams) 59.9
48.6 38.9 147.4 M C1 (High yield) 45.2 37.4 39.5 122.1 122.1/35.3
147.4/46.1 (wt/plant in grams) % Hybridity (56.9) (72.2) (74.8)
F(A)(wt/pl) in grams 17.4 15.3 13.4 46.1 M C2 (Low yield) 12.9 11.8
10.6 35.3 (wt/plant in grams) Avg. Female seed wt. 38.7 32.0 26.2
Ave. Male seed wt. 29.1 24.6 25.1 % Hybridity (62.5) (73.1)
(79.9)
[0043] In one preferred embodiment of the method of the invention
to produce a hybrid seed product, selection of a cross having a
high female to pollenizer seed yield differential provides for an
increase in the percentage of hybridity of the hybrid seed product.
The teachings of the present invention demonstrates that seed yield
of female plants varies in response to different males at different
female to pollenizer ratios. Table 5 illustrates results of seed
yield for a female line crossed with a high seed yield pollenizer
and a low seed yield pollenizer at three different female to
pollenizer ratios. Thus, greater levels of hybridity can be
obtained by selection for a cross where the pollenizer lines and
female lines have a high female to pollenizer seed yield
differential. In the preferred embodiment, the methods for
producing a hybrid seed product of the instant invention include
selection of a female line and a pollenizer line to provide for a
high female to pollenizer seed yield differential.
[0044] The present invention further illustrates, as shown in Table
5, that female seed yield has a positive correlation with
decreasing female to pollenizer ratios. Total seed yield also has a
positive correlation with decreasing female to pollenizer ratios.
In contrast, the percentage of hybridity has a positive correlation
with increasing female to pollenizer ratios. Female seed yield is
also affected by pollenizer seed yield, with higher pollenizer seed
yield positively correlated to higher female seed yield.
TABLE-US-00006 TABLE 6 Female1 with Male1 Seed Yield Decrease and
Hybridity Increase as Female to Male (Pollenizer) Ratios Changed
from 1:1 Ratio to 2:1 Ratio and 3:1 Ratio Female:Male 1:1 2:1 3:1
F1 (wt/pl) grams 59.9 48.6 38.9 % reduction from 1:1 18.9 35.1 seed
production level M1 (wt/pl) grams 45.2 37.4 39.5 % reduction from
1:1 17.3 12.6 seed production level Total (Female + Male) 105.1
86.0 78.4 % reduction from 1:1 18.2 25.4 seed production level F:M
seed yield differential 1.33 1.30 0.98 % Hybridity 57.0 72.2
74.7
TABLE-US-00007 TABLE 7 Female1 with Male2 Seed Yield Decrease and
Hybridity Increase as Female to Male (Pollenizer) Ratios Changed
from 1:1 Ratio to 2:1 Ratio and 3:1 Ratio Female:Male 1:1 2:1 3:1
F1 (wt/pl) grams 17.4 15.3 13.4 % reduction from 1:1 12.1 23.0 seed
production level M2 (wt/pl) grams 12.9 11.8 10.6 % reduction from
1:1 8.5 17.8 seed production level Total (Female + Male) 30.3 27.1
24.0 % reduction from 1:1 10.6 20.8 seed production level F:M seed
yield differential 1.35 1.30 1.26 % Hybridity 62.5 73.1 79.9
TABLE-US-00008 TABLE 8 Overall changes in Seed Yield Decrease and
Hybridity Increase as Female to Male (Pollenizer) Ratios Increases
from 1:1 Ratio to 2:1 Ratio and 3:1 Ratio Female:Male 1:1 2:1 3:1
F1 (wt/pl) grams 59.9 48.6 38.9 Change 100.0 81.1 64.9 M1 (wt/pl)
grams 45.2 37.4 39.5 Change 100.0 82.7 87.4 Total (Female + Male)
105.1 86.0 78.4 Change 100.0 81.8 74.6 F:M seed yield differential
1.33 1.30 .99 % Hybridity 56.9 72.2 74.8
[0045] Tables 6, 7 and 8 show data for a female line crossed with
two pollenizer lines using three different female to pollenizer
ratios. Table 6 shows the female line crossed with a first
pollenizer line (Male1) at three female to pollenizer ratios. Table
7 shows the same female line crossed with, a second pollenizer line
(Male2) at three female to pollenizer ratios. Data obtained from
experimentation according to the teaching of the present invention
demonstrates an average hybridity level of 71.8% for the cross with
the first pollenizer line and an average hybridity level of 67.8%.
for the cross with the second pollenizer line. Thus, according to
the teachings of the present invention, percentage of hybridity
will vary from cross to cross.
[0046] This experimentation further shows differences in response
to seed yield and percentage of hybridity from cross to cross.
Table 6 shows that for the first pollenizer (Male1), seed yield
increased by 34% and 10% when female to pollenizer ratio changed
from 3:1 to 1:1 and 3:1 to 2:1, respectively. Percentage of
hybridity, conversely, increased by 31% and 27% when female to
pollenizer ratios changed from 1:1 to 3:1 and 1:1 to 2:1,
respectively. Table 7 shows that for the second pollenizer (Male2),
seed yield increased by 26% and 13% when female to pollenizer ratio
changed from 3:1 to 1:1 and 3:1 to 2:1, respectively. The
percentage of hybridity increased by 28% and 17%, conversely, when
female to pollenizer ratios changed from 1:1 to 3:1 and 1:1 to 2:1,
respectively. Thus, according to the teachings of the present
invention, in the preferred embodiment of the invention for
producing a hybrid seed product, each cross is tested for
responsiveness to female to pollenizer ratio for seed yield and
percentage of hybridity. In the preferred embodiment, a cross is
selected that maintains at least two-thirds of female seed yield
when increasing ratio of female plants to pollenizer plants from
1:1 to 3:1 under substantially similar environmental conditions.
Alternatively, for some varieties, the method for producing a
hybrid seed product can make use of a generic formula to estimate
responsiveness to female to pollenizer ratio tor seed yield and
percentage of hybridity. One way of establishing a generic curve is
by estimating a curve from multiple crosses of other varieties.
FIG. 2A shows one example embodiment of a curve estimated from
several female-pollenizer crosses.
TABLE-US-00009 TABLE 9 The effect on hybrid seed yield and
percentage of hybridity by changing female to pollenizer ratios.
Female to Pollenizer Ratios. Female 70:30 75:25 80:20 F1 SEEDYIELD
345 298 218 % CHANGE 100 86 63 HYBRIDITY 87.5 93.5 90.5 % CHANGE
100 107 103 F2 SEEDYIELD 285 357 231 % CHANGE 100 125 81 HYBRIDITY
88.5 84.5 87.5 % CHANGE 100 96 99 F3 SEEDYIELD 340 174 193 % CHANGE
100 51 57 HYBRIDITY 84.5 88 91 % CHANGE 100 104 108 F4 SEEDYIELD
344 285 252 % CHANGE 100 83 73 HYBRIDITY 83.5 88.00 89.00 % CHANGE
100 105 107 OVER ALL SEEDYIELD 100 86 69 CHANGE HYBRIDITY 100 103
104
[0047] Table 9 summarizes further experimental seed yield and
percentage of hybridity tor multiple female and pollenizer crosses
at different female to pollenizer ratios. In the most preferred
embodiment, production of a hybrid seed product includes a
statistical analysis on data for a selected cross in order to
determine the effect of different female to pollenizer ratios on
seed yield. In one example embodiment, the statistical analysis
comprises a regression analysis. FIG. 2B shows a number of
regression curves for different female-pollenizer crosses.
[0048] As shown in Table 9, according to the teachings of the
present invention, changing female to pollenizer ratios from a
70:30 ratio to a 75:25 ratio or a 70:30 ratio to an 80:20 ratio
greatly impacts hybrid seed yield. Seed yield dropped an average of
31% from a 70:30 ratio to 80:20 ratio and dropped an average of 16%
from a 70:30 ratio to a 75:25 ratio, respectively, when Male3 was
crossed with four different female lines. Average percentage of
hybridity increased 3% when female to pollenizer ratios changed
from a 70:30 ratio to a 75:25 ratio and increased 4% when going
from a 70:30 ratio to an 80:20 ratio, respectively.
[0049] While a hybrid seed product having a high percentage of
hybridity is desirable for agronomic qualities and for
certification; on the other hand, the cost of producing a hybrid
seed product increases with decreasing seed yield. The methods for
producing a hybrid seed product of the present invention,
therefore, provides methods for determining a female to pollenizer
ratio providing for maximum seed yield at a predetermined level of
hybridity of the hybrid seed product.
TABLE-US-00010 TABLE 10 Hybrid Seed Yield and Percentage of
Hybridity are Different with Each Specific Combination of Female
Line and Pollenizer at Different Female to Pollenizer Ratios Female
seed % Female seed F:M % of Male. Hybridity yield Change % F1:M1
0.7:1.0 .sup. 78.7/100.0 44.0 3:1 61.6/100 64.9 Female ratio
change. (0.7:1.00 to 3:1) (78.3%) F2:M2 1.1:1.0 .sup. 92.3/100.0
48.0 3.2:1.0 65.1/100 75.3 Female ratio change. (1.1:1.0 to
3.2:1.0) (70.5) F3:M3 1:1 141.0 58.5 2.5:1.sup. 97.8/100 71.0
Female ratio change. (1:1 to 2.5:1) (69.4) F1:M4 0.8:1.0 101.4/100
50.3 3.0:1.0 115.1/100 77.5 Female ratio change (0.8:1.0 to 3:1)
(113.5) 5 2.3:1.0 86.8/100 66.6 3.0:1.0 100/100 75.0 Female ratio
change (2.3:1.0 to 3:1) (115.2)
[0050] Table 10 summarizes experimentation according to the
teachings of the present invention to determine varying effects of
female to pollenizer ratios on different crosses. In this example
embodiment, Female1 (F1), Female2 (F2) and Female3 (F3) seed yield
decreased at higher female to pollenizer ratios when pollinated by
Pollenizer1 (M1) and Pollenizer2 (M2). Female4 (F4) and Female5
(F5), on the other hand, did not show seed yield decrease at higher
female to male ratios when pollinated by Pollenizer4 (M4) and
Pollenizer5 (M5). With respect to the lines tested and reported on
Table 9, (A1XB1) X (A2XR (Restorer)) cross demonstrates both higher
hybrid seed yield and higher percentage of hybridity.
[0051] In the preferred embodiment of the invention, the data
analysis 40 comprises a regression analysis of percentage of
hybridity versus female to pollenizer ratio to establish a
regression curve for each individual cross. In this embodiment of
the present invention, selection of lines for hybrid seed
production employs the resulting regression curves for each cross
under consideration in order to select a cross for high seed yield
during hybrid seed production. FIG. 2B shows a regression curve
showing percentage hybridity versus female to pollenizer ratio for
multiple crosses. In this example embodiment, the regression
analysis shows:
y=18.836ln(x)+55.128
[0052] where: y=percentage of hybridity [0053] x=female to
pollenizer ratio.
[0054] In this embodiment of the invention to produce a hybrid seed
product, the regression analysis allows for determination of a
female to pollenizer ratio necessary to achieve a targeted
percentage of hybridity. In one example embodiment, selection of
the female to pollenizer ratio employs the regression analysis to
target a 75% hybridity level to achieve certification standards for
a hybrid variety. In an alternative embodiment, selection of the
female to pollenizer ratio employs the regression analysis to
target a 95% hybridity level.
[0055] However, in many female to pollenizer crosses, seed yield
dramatically decreases with increasing female to pollenizer ratios.
Agronomic or other considerations often suggest a cross having a
dramatic decrease in seed yield with increasing female to
pollenizer ratios. Therefore, the present invention provides a
means for post-production increase of percentage of hybridity in a
hybrid seed product.
[0056] In the preferred embodiment of the invention, the method for
production of a hybrid seed product increases the percentage of
hybridity of a hybrid seed product by removing non-hybrid seed. In
one example embodiment, removal of non-hybrid seed makes use of
seed size differential between hybrid seed produced by female
plants and non-hybrid seed produced by pollenizer plants. In this
preferred embodiment, selection of a female line and selection of a
pollenizer line results in hybrid seed having an average size
larger than non-hybrid seed. In an alternate embodiment, selection
of a female line and selection of a pollenizer line results in
hybrid seed having an average size smaller than non-hybrid
seed.
[0057] Testing according to the preferred teachings of the
invention demonstrates selection of a female line and a pollenizer
line can provide a cross having a high correlation (r=0.9) between
female parental lines and their first generation progeny with
respect to seed size. Thus, in the preferred embodiment, the
methods for producing a hybrid seed product employs selection for
hybrid seed having a seed size distribution skewed to larger seed
sizes and non-hybrid seed having a seed size distribution skewed to
smaller seed sizes. Sifting production seed to screen out seed
smaller than a selected seed size increases the percentage of
hybrid seed for the hybrid seed product. In one example embodiment,
a sieve screens out seed smaller than a selected seed size to
increase the percentage of hybridity of the hybrid seed product. In
one example embodiment, shown in Table 14, using a 1/19'' sieve
provides a gain to 75% hybridity when production of hybrid seed is
done using a 65:35 ratio of female plants to pollenizer plants. As
those skilled in the art will recognize, different sieve sizes can
be employed to accomplish different increases in percentage of
hybridity.
TABLE-US-00011 TABLE 12 Seed Size Distributions of a Female Line, 3
Pollenizers and a Hybrid (A1 .times. B1) .times. (C1 + C2 + C3)
1/14 1/15 1/16 1/17 1/18 1/19 1/20 1/21 1/22 1/23 REMAIN MEAN H1
(hybrid) 0.49 14.99 25.22 25.72 20.97 8.47 0.35 3.54 0.22 0.01 0.00
5.93 F1 (Female 1) 1.17 18.60 23.73 26.24 18.11 7.54 0.22 3.72 0.28
0.35 0.00 5.96 C1 (Pollenizer 1) 0.73 1.46 4.47 15.56 29.18 21.97
2.09 21.49 2.58 0.19 0.29 5.37 C2 (Pollenizer 2) 0.56 2.08 7.21
21.01 21.86 24.32 1.91 19.14 1.80 0.12 0.00 5.45 C3 (Pollenizer 3)
0.80 10.64 21.78 27.68 20.08 10.16 0.58 7.17 0.83 0.26 0.01 5.82
Mean of C1 + C2 + C3 0.70 4.73 11.16 21.41 23.71 18.82 1.53 15.93
1.74 0.19 0.10 1,000 SEED WEIGHT H1 = (A1 .times. B1) .times. (C1,
C2, C3) 2.480 GRAMS F1 = A1 .times. B1 2.365 GRAMS C1 2.200 GRAMS
C2 2.339 GRAMS C3 2.250 GRAMS
TABLE-US-00012 TABLE 13 Seed Size Distribution of a Female Line, 3
Pollenizers a Hybrid Using a 1/19'' sieve to 1/23'' sieve Entry
1/19 1/20 1/21 1/22 1/23 Hybrid (A1XB1) 12.59% 4.12% 3.77% 0.23%
0.01% XC1, C2, C3) Female (AXB) 11.92% 4.58% 4.36% 0.63% 0.35%
Pollenizer (C1) 47.37% 26.93% 24.87% 2.87% 0.23% Pollenizer (C2)
46.26% 23.38% 21.47% 1.96% 0.12% Pollenizer (C3) 18.54% 8.90% 8.32%
1.11% 0.27% Pollenizer Average 37.39% 19.74% 18.22% 1.98% 0.21%
[0058] As shown in Table 12, seed size differential varies for each
crossing of different female lines and pollenizer lines. Table 12
shows a seed size distribution for a hybrid line, a female line,
and three pollenizer lines. Table 13 shows the percentage of seed
screened out using five different sieve sizes, from 1/19 to
1/23.
[0059] According to the teachings of the present invention, seed
size distributions vary for each different cross; therefore,
increases in hybridity level also vary with each different cross
between different female lines and different pollenizer lines.
Because seed size is a inheritable trait, in the preferred
embodiment of the present invention for producing a hybrid seed
product, selection of female and pollenizer lines provides for seed
size differential between hybrid and non-hybrid seed. Environmental
conditions, such as drought and poor soil conditions, also affect
seed size. Thus, determination of seed size distribution according
to the teachings of the present invention, should be done in the
same field to control for environmental variation. As the seed size
distribution differential between female line and pollenizer line
increases, the environmental effect on the use of screening to
remove non-hybrid seed will be reduced. Experimentation according
to the teaching of the present invention, and as shown in Table 14,
illustrates the increase in hybridity levels achieved by straining
out a selected seed size for crosses between one female line and
three pollenizer lines. In particular, Tables 14a and 14b
illustrate gains in hybridity level for a 1/19'' sieve and a 1/21''
size sieve for the selected crosses.
Percentage of Hybridity Gained by Applying 1/19'' Sieve and 1/21''
Sieve to Screen Out a Higher Percentage of Non-Hybrid Seed
(Pollenizer C.sub.1, C.sub.2, C.sub.3)
TABLE-US-00013 [0060] TABLE 14a AMT. SEED SCREEN FEMALE: % *%
HYBRID PARENTS OUT FOR 1/19 SIZE POLLENIZER HYBRIDITY FOR FEMALE
POLLENIZER FEMALE POLLENIZER RATIO INC. PRODUCT A1 .times. B1 C1
11.92% 47.37% 65:35 10.7 75.7 C2 46.26% 10.3 75.3 C3 18.54% 1.8
66.8 C1 70:30 9.6 79.6 C2 9.3 79.3 C3 1.6 71.6 C1 75:25 8.4 83.4 C2
8.1 83.1 C3 1.4 76.4 C1 80:20 7.0 87.0 C2 6.8 86.8 C3 1.2 81.2
TABLE-US-00014 TABLE 14b AMT. SEED SCREEN FEMALE: % % HYBRID
PARENTS OUT FOR 1/21 SIZE POLLENIZER HYBRIDITY FOR FEMALE
POLLENIZER FEMALE POLLENIZER RATIO INC. PRODUCT A1 .times. B1 C1
4.36% 24.87% 65:35 5.3 70.3 C2 21.47% 4.3 69.3 C3 8.32% 1.0 66.0 C1
70:30 4.8 74.8 C2 4.0 74.0 C3 0.9 70.9 C1 75:25 4.3 79.3 C2 3.5
78.5 C3 0.8 75.8 C1 80:20 3.6 83.6 C2 3.0 83.0 C3 0.7 80.7
[0061] In alternate embodiments of the invention, the method for
post-production increase of percentage of hybridity can employ
differences in seed density, seed weight or seed coat color. In one
example embodiment, selection of a female line and a pollenizer
line may provide for non-hybrid seed having a lesser density than
hybrid seed. In this example embodiment, the teachings of the
present invention provides for removal of non-hybrid seed according
to seed density or seed weight. A gravity table or rice roller can
separate hybrid and non-hybrid seed in this embodiment of the
invention. In an alternate embodiment employing differences in seed
color between hybrid seed and non-hybrid seed, a seed coat color
sorter removes non-hybrid seed. Those skilled in the art will
recognize that other methods of removing non-hybrid seed to
increase the percentage of hybridity of a hybrid seed product lie
within the spirit and scope of the invention. For example,
morphological traits such as seed size, seed color, seed weight,
leaf shape, leaf size, root color, stem color, flower color root
structure, plant height and fall growth habit or a genetic marker
can be used to distinguish hybrid seed from non-hybrid seed.
[0062] Therefore, in the preferred embodiment of the invention to
produce certified seed, the regression curve of a targeted hybrid
product provides for selection of a female to pollenizer ratio,
that when combined with a post-production increase in hybridity
level, achieves maximum yield at a hybridity level of at least
75%.
[0063] In the preferred embodiment of the invention to produce a
hybrid seed product, test plots provide for determination of
percentage of hybridity at different female to pollenizer ratios.
Regression analysis of percentage of hybridity versus female to
pollenizer ratio then provides a regression curve for each
individual cross to allow for production of a hybrid product having
a preselected percentage of hybridity. In other aspects of the
method of the invention, the test plots further provide for
determination of the seed size distribution of selected female
lines, male lines and hybrid lines. The test plots also provide for
determination of the 1,000 seed weight of each line and the P.P.I.
of the female lines.
[0064] In one example embodiment of the invention to produce a
hybrid seed product, the following types of test plots are
employed.
[0065] A "Check" plot has female seed and pollenizer seed planted
in the same plot. The check plot includes two pollenizer rows, a
skipped row, four female rows, a skipped row, and two pollenizer
rows. This design of the "Check" plot allows the female rows and
pollenizer rows to be harvested separately. After flowering,
analysis of the female plants provides for determination of the
P.P.I. After harvesting, analysts of the seed from the female
plants provides determination of seed size distribution and 1000
seed weight. After harvesting the pollenizer plants, analysis of
the seed from pollenizer plants provides determination of seed size
distribution and 1000 seed weight. Analysis also determines data
for the seed yield of female plants and pollenizer plants.
[0066] A female to pollenizer plot designed to have a 1:1 ratio.
The female and pollenizer plants are intermingled and distributed
randomly.
[0067] A female to pollenizer plot designed to have a 2:1 ratio.
The female and pollenizer plants are intermingled and distributed
randomly.
[0068] A female to pollenizer plot designed to have a 3:1 ratio.
The female and pollenizer plants are intermingled and distributed
randomly.
[0069] A female to pollenizer plot designed to have a 4:1 ratio.
The female and pollenizer plants are intermingled and distributed
randomly.
[0070] In one preferred embodiment, the test plots are planted
using a complete random block design with four to eight
replications for each female to pollenizer ratio. In this
embodiment, the test plots are six row plots, with each row fifty
to one hundred feet long. Data collection is performed on the
middle two to four rows, with the outside rows serving as borders.
The outside border rows serve as a guard and a barrier to reduce
contamination from neighboring plots.
[0071] In this example embodiment of the invention, data collection
follows the following procedures. The number of seed produced by
female plants can be determined using the equation:
x=female seed weight/weight of one thousand female seed,
[0072] where x=the mean number of seed produced by a female
plant.
[0073] The number of seed produced by pollenizer plants can be
determined using the equation:
y=pollenizer seed weight/weight of one thousand pollenizer
seed,
[0074] where y=the mean number of seed produced by a pollenizer
plant.
[0075] The percentage of female seed can be then determined as:
% female seed=x/(x+y).
[0076] In the preferred embodiment of the invention, determination
of percentage of hybridity takes selfing and sibbing in the female
line into account. The adjusted percentage of hybridity corrects
for these factors by subtracting non-hybrid seed produced from
female plants. The ASOCA publication of hybrid alfalfa
certification provides the correction factor for non-hybrid seed
produced from female plants from sibbing and selfing. The
correction factor is equal to P.P.I..times.0.595, with the P.P.I.
(Pollen production index) defined as follows.
[0077] 1. Male Sterile Plants (MS) P.P.I.=0 [0078] No visible
pollen can be observed with the naked eye when flower is tripped
with a black knife blade.
[0079] 2. Partial Male Sterile Plants (PMS) P.P.I.=0.1 [0080] A
trace of pollen can be observed with the naked eye when flower is
tripped with a black knife blade.
[0081] 3. Partial Fertile Plant (PF) P.P.I.=0.6 [0082] Less than
normal amount of pollen can be observed with the naked eye when
flower is tripped with a black knife blade.
[0083] 4. Fertile Plant (P) P.P.I.=1.0 [0084] Normal amounts of
pollen can be observed when flower is tripped with a black knife
blade.
[0085] In this example embodiment of the invention, the pollen
production index (P.P.I.) is determined by sampling 200 female
plants from the female rows of the "check" plots. The percentage of
hybridity can then be determined by the equation:
% hybridity=% female seed-% non-hybrid female seed,
[0086] where the % non-hybrid female seed is P.P.I..times.0.595
[0087] In one preferred embodiment of the invention, the methods
for producing a hybrid seed product determines the percentage of
hybridity for each female to male ratio of 1:1, 2:1, 3:1, and
4:1,
[0088] This method for determining the percentage of hybridity
requires analysis of seed produced by female plants apart from
analysis of seed produced by pollenizer plants. Harvesting female
plants separately from pollenizer plants is one way of obtaining
female seed apart from pollenizer seed. However, separate
harvesting of female plants from pollenizer plants is difficult
under current hybrid seed production methods.
[0089] In one method for hybrid seed production, female plants are
crossed with pollenizer plants by planting female plants and
pollenizer plants in separate rows of plants, with each row
separated by a width of twenty to forty inches. Each group of rows
contain either pollenizer plants or female plants, such as shown in
U.S. Pat. No. 4,045,912, herein incorporated by reference. This
allows female plants and pollenizer plants to be harvested
separately according to rows. However, this method involves a large
physical separation of female and pollenizer plants, resulting in
lower production of hybrid seed.
[0090] Female and pollenizer plants may also be intermingled in the
same row in the same test plot. This intermingling increases hybrid
seed production from the female plants. However, intermingling also
causes difficulty in distinguishing the female plants from the
pollenizer plants. To obtain data on hybrid seed production, seed
from female plants must be distinguished from seed from pollenizer
plants. Distinguishing between female plants and pollenizer plants
is a highly labor intensive, difficult and expensive process. U.S.
Pat. No. 4,045,912 provides one example of such a process, where
female and pollenizer plants are germinated separately, and then
hand planted and tagged in a test plot. This process becomes
untenable in larger applications. Thus, the present invention
provides methods for distinguishing female plants from pollenizer
plants suitable for large-scale applications, such as large test
plots.
[0091] The preferred embodiment of the invention for producing a
hybrid seed product provides for methods for distinguishing female
plants from pollenizer plants by planting female lines and
pollenizer lines in subrows. The subrow methods of the invention
further allow for separate harvesting of female plants and
pollenizer plants. During planting, female plants and pollenizer
plants are offset from the center of a row on opposite sides to
distinguish the female plants from the pollenizer plants.
[0092] FIG. 3 shows an example embodiment of the subrow planting
method of the invention. The subrow planting method provides for
planting both female and pollenizer plants in a single row 200,
with female seed planted two to four inches from the center 210 of
the row 200 on a first subrow 220, and pollenizer seed are planted
one to eight inches from the center 210 of the row 200 in a second
subrow 230 on the opposite side. Thus, in this example embodiment,
planting according to the subrow methods provides female plants 240
offset to one side in the first subrow 220 and a pollenizer plants
250 in the second subrow 220 offset to the other side. This single
row 200 shows an embodiment having a female to pollenizer ratio of
2:1. A second single row 260 shows an embodiment having a female to
pollenizer ratio of 3:1. A third single row 270 shows a female to
pollenizer ratio of 4:1. In one preferred embodiment of the
invention, an onion planter may be used to plant female seed and
pollenizer seed in their respective subrows. Those skilled in the
art will recognize that other methods of planting the female seed
and the pollenizer seed in their respective subrows lies within the
spirit and scope of the invention.
[0093] In other aspects of the subrow methods of the invention, the
subrows 220, 230 can advantageously provide for reduction of
competition between female plants and male plants by increasing
physical separation of the female plants from the pollenizer plants
while still allowing intermingling. In cases where one line may be
more vigorous than another, the increased separation between plants
may allow slower growing plants a better opportunity to grow
without being crowded out by other plants. Because of the small
seed size of some field crops, such as alfalfa, during planting,
some seeds are not spaced with sufficient room to grow, allowing a
faster growing plant to crowd out a slower developing plant,
particularly in subsequent years of perennial crops. Subrows
provide for increased separation between female plants and
pollenizer plants that reduces competition between the female
plants and pollenizer plants, maintaining a more consistent female
to pollenizer ratio. As the female to pollenizer ratio is selected
to optimize production of the hybrid seed product, in this aspect
of the present invention, planting in subrows in the production
field maintains optimal production from year to year.
[0094] In another aspect of the subrow methods of the invention,
subrows 220, 230 allow a breeder to quickly determine the actual
female to pollenizer ratio. At a seedling stage, the subrows 220,
230 allow the number of female plants 240 and the number of
pollenizer plants 250 to be quickly counted. In subsequent years
after planting for perennial plants, a new female to pollenizer
ratio needs to be determined from year to year, as plants may die
over the winter. In this instance, subrows 220, 230 facilitate
quick determination of female to pollenizer ratio through visual
inspection.
[0095] In another aspect of the invention, subrows 220, 230
facilitate separate harvesting of female plants 240 and pollenizer
250 by harvesting according to female subrows and pollenizer
subrows. Separate harvesting allows for determination of percentage
of hybridity by sampling before harvest.
[0096] In an alternate embodiment of the invention, morphological
markers, such as seed size, seed color, seed weight, leaf shape,
leaf size, root color, stem color, flower color root structure,
plant height and fall growth habit flower colors, can be used to
distinguish female plants from pollenizer plants in the test plots.
Those skilled in the art will recognize that other methods of
distinguishing female plants from pollenizer plants can be employed
without departing from the spirit and scope of the invention.
[0097] The preferred form of the invention further includes
verification of percentage of hybridity of the hybrid seed product
140. These methods involve progeny testing and are employed after
production of the hybrid seed product in the production field.
Verification of percentage of hybridity can be used to help ensure
a hybrid seed product achieves predetermined hybridity goals. In
the preferred embodiment, a characteristic of the hybrid seed that
has a distinct difference from non-hybrid seed is used in the
verification process. However, as those skilled in the art will
understand, other methods for verifying percentage of hybridity can
be employed without departing from the spirit or scope of the
invention.
[0098] In one preferred form, verifying percentage of hybridity
begins with choosing a row and sampling 200-1000 female plants and
200-1000 pollenizer plants. Data is collected to determine the seed
yield of the female plants and the seed yield of the pollenizer
plants. The female to pollenizer ratio is determined by counting
number of female plants to number of pollenizer plants for 200 feet
in a chosen row for two to five replications in a hybrid seed
production field. The subrow system can be employed to aid tracking
of female plants versus the pollenizer plants to distinguish the
plants in the commercial hybrid seed production field.
[0099] Analysis on the gathered data then provides an estimated
percentage of hybridity. Because of the variability inherent in
biological systems, the analysis provides an estimate rather than
an exact determination. The analysis can be performed according to
the following procedures:
[0100] 1. From the sampled plants, determine the average female
seed yield (X) to pollenizer seed yield (Y) per plant and determine
average number of seed produced from female (hybrid) plants to
number of seed produced from pollenizer plants as X:Y.
[0101] 2. Determine number of female plants to number of pollenizer
plants in a commercial hybrid seed production field by measuring
the number of female plants and number of pollenizer plants in 100
feet of a randomly selected row with two to five replications. The
proportion of female plants to the number of pollenizer plants is
A:B.
[0102] 3. The percentage of hybridity is computed according to:
% Hybridity = ( A .times. X ) ( A .times. X ) + ( B .times. Y )
.times. 100 % - CF ##EQU00001##
[0103] where CF is a correction factor to account for non-hybrid
seed produced from female plants by selfing, and is equal to the
P.P.I. (female plants).times.0.595.
[0104] Harvesting the center two rows of pollenizer line from the
pollenizer test strip and harvesting the center two rows of the
female line from the female test strip separately provides for data
on seed size distribution of the pollenizer seed and hybrid seed.
The commercial hybrid production field is harvested in bulk.
[0105] In another example embodiment, seed size distribution
differential between hybrid seed and pollenizer seed provides data
for verifying percentage of hybridity. In this method, four to six
rows of 200-1000 feet of plants from a selected pollenizer line are
planted as a pollenizer test strip and four to six rows of 200-1000
feet of plants from a selected female line are planted as a female
test strip in the same commercial hybrid seed production field.
[0106] Analysis of the pollenizer seed provides the pollenizer seed
size distribution. Likewise, analysis of the female seed, provides
the female seed size distribution, and analysis of the
bulk-harvested seed provides the seed size distribution of the
commercial hybrid production field. By way of illustration and not
limitation, the following shows an example with the seed size
distribution determined for each increment from 1/10 inch to 1/25
inch. In this example, samples having the same weight are used to
determine the seed size distributions. The following procedure
provides an estimate of the percentage of hybridity.
[0107] 1. Perform a T-test for seed size distribution of pollenizer
seed, hybrid seed and commercial experimental hybrid seed
(female:pollenizer ratio block used by commercial hybrid seed
production field) harvested from a test plot, and comparing with
pollenizer seed, hybrid seed and commercial hybrid seed harvested
from a commercial hybrid seed production field.
[0108] 2. If the T-test is not significant, then the percentage of
hybrid seed in the overlap area of the seed size distribution in
the test plot would be same as overlap area of the seed size
distribution in the commercial hybrid production seed field.
[0109] The % hybridity in the commercial hybrid production seed
field can then be determined by:
% Hybridity = X 1 + Y 2 X 1 + Y 2 + Q 2 + R 1 .times. 100 % - CF
##EQU00002##
Where:
[0110] X=Hybrid seed weight of non-overlap area.
[0111] X1=X/1,000 seed weight of non-overlap area.
[0112] Y=Hybrid seed weight and pollenizer seed weight of overlap
area.
[0113] Y1=Y.times.% Hybrid seed overlap area (from test plots).
[0114] Y2=Y1/1,000 seed weight.
[0115] Q=Pollenizer seed weight of overlap area.
[0116] Q1=Y.times.% Pollenizer seed of overlap area.
[0117] Q2=Q/1,000 seed weight.
[0118] R=Pollenizer seed weight of non-overlap area.
[0119] R1=R/1,000 seed weight.
[0120] CF=P.P.I. (female plants).times.0.595.
[0121] In an alternate embodiment, seed size distribution can be
used to verify percentage of hybridity using the proportion of seed
within a selected seed size range. The selected seed size range
includes an area of overlap in the seed size distribution between
the seed size distribution of the female seed and pollenizer seed.
The percentage of hybridity can be estimated as:
% Hybridity = Z - Y X - Y .times. 100 % - CF ##EQU00003##
[0122] where:
[0123] X=percentage of female seed fatting into the selected seed
size range;
[0124] Y=percentage of pollenizer seed falling into the selected
seed size range;
[0125] Z=percentage of production seed falling into the selected
seed size range; and
[0126] CF=correction factor to account for non-hybrid seed produced
from female plants by selfing, and is equal to the P.P.I. (female
plants).times.0.595.
[0127] As those skilled in the art will recognize, other seed
characteristics where the female seed has a distinct difference
from the pollenizer seed can be used to estimate the percentage of
hybridity. For example, in an alternate embodiment, the mean seed
size of the seed sampled from the pollenizer plants, the female
plants and the production field can be used to estimate percentage
of hybridity. Using this method, percentage of hybridity can be
estimated as:
% Hybridity = Z - Y X - Y .times. 100 % - CF ##EQU00004##
[0128] where:
[0129] X=mean female seed size;
[0130] Y=mean pollenizer seed size;
[0131] Z=mean production seed size; and
[0132] CP=correction factor to account for non-hybrid seed produced
from female plants by selling and sibbing, equal to the P.P.I.
(female plants).times.0.595,
[0133] In another example embodiment, the methods for verification,
of percentage of hybridity employ the pollen production index
(P.P.I.) of hybrid seed and the commercial hybrid seed product,
which is a mixture of hybrid and pollenizer seed. In this
embodiment of the invention employing the P.P.I., verification of
percentage of hybridity begins with crossing a selected female line
with a selected pollenizer line and harvesting the seed produced by
the cross in a test plot and production field. The harvested seed
is planted and P.P.I. data is gathered. Further analysis for
verification of hybridity proceeds in one of two cases, depending
on whether the P.P.I. of the hybrid seed production field is larger
than the P.P.I. of the test plot hybrid seed or vice versa.
CASE 1
[0134] If the P.P.I. of the hybrid seed production field is larger
than the P.P.I. of the test plot hybrid seed, the following formula
will be used to calculate the percentage of hybridity.
[0135] The seed from a hybrid seed production field contain both
hybrid seed (female X pollenizer) and non-hybrid seed (pollenizer).
The seed from test plot of f:p=1:0 contains only hybrid seed.
[0136] a. % of hybridity=1-P.P.I. (Production seed)/1-P.P.I. (Test
plot hybrid seed).times.100-CF
[0137] b. Sample size of growouts=200-2000 plants from each
population (production seed and test plot hybrid seed) at the same
environment and same location.
CASE 2
[0138] If the P.P.I. from the hybrid seed production field is
similar to the P.P.I. of the test plot hybrid seed, then seed size
distribution differential between hybrid seed and pollenizer seed
or some morphological traits or molecular markers that can
differentiate hybrid seed from non-hybrid seed from grow outs need
to be used to verify the percentage of hybridity.
[0139] As the invention disclosed herein may be embodied in other
specific forms without departing from the spirit or general
characteristics thereof, some of which forms have been indicated,
the embodiments described herein are to be considered in all
respects illustrative and not restrictive. The scope of the
invention is to be indicated by the appended claims, rather than by
the foregoing description, and all changes which come within the
meaning and range of equivalency of the claims are intended to be
embraced therein.
* * * * *