AU734755B2 - A method to identify and breed corn with increased kernel oil concentration - Google Patents

A method to identify and breed corn with increased kernel oil concentration Download PDF

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AU734755B2
AU734755B2 AU65751/98A AU6575198A AU734755B2 AU 734755 B2 AU734755 B2 AU 734755B2 AU 65751/98 A AU65751/98 A AU 65751/98A AU 6575198 A AU6575198 A AU 6575198A AU 734755 B2 AU734755 B2 AU 734755B2
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Robert Stefan Reiter
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Description

WO 98/42870 PCT/US98/05550
TITLE
A METHOD TO IDENTIFY AND BREED CORN WITH INCREASED KERNEL OIL CONCENTRATION FIELD OF INVENTION The inveition is in the fields of plant breeding and molecular biology.
More specifically, the invention relates to the identification of corn loci conferring increased kernel oil concentration using genetic markers and the use of genetic markers as an aid to the identification and breeding of corn with increased kernel oil concentration.
BACKGROUND OF INVENTION Corn is a major crop used as a human food source, an animal feed, and as a source of carbohydrate, oil, protein, and fiber. It is principally used as an energy source in animal feeds, or as a raw material for the recovery of starch, protein feed fractions, fiber, flaking grits, flour, and oil.
Most commercial corn produced throughout the United States is produced from hybrid seed. The production of corn hybrids requires the development of elite corn inbreds that upon intermating produce agronomically superior hybrids.
During the development of corn inbreds, plant breeders select for a number of different traits affecting agronomic performance. These traits include but are not limited to stalk strength, lodging, disease resistance, grain moisture and grain yield. Agronomic traits tend to be quantitatively measured with continuous rather than discrete distributions. It is theorized that quantitative traits are controlled by several genes with small and generally equivalent effects. Further, the observed phenotype is due partially to this genetic component and an environmental component.
The heritability of a trait is defined in the broad sense as the ratio of the genetic variance to the total phenotypic variance. Many agronomic traits display low heritability; the performance of parent plants is a poor predictor of offspring performance. Thus, traits with low heritability have small genetic variance components in comparison with observed variation. The impact on the plant breeder is that in breeding populations, the value of a plant's genetic composition is difficult to determine from agronomic trait measurements. In an attempt to maximize their discriminative abilities, breeders collect multiple measurements both from individuals related by descent and from many environments. This strategy is resource intensive because it involves the use of extensive trialing to make even small gains in plant improvement. This, coupled with the fact that improved corn lines are selected for multiple traits simultaneously, makes the development of superior corn inbreds both a timeconsuming and an expensive labor.
WO 98/42870 PCT/US98/05550 The addition of novel traits in a corn breeding program imposes an additional burden upon the plant breeder. Depending upon the genetic complexity of the novel trait single gene versus many genes), a significant increase in time and effort is required to produce elite lines containing novel traits. One such trait is kernel oil concentration.
Corn with increased kernel oil concentration is important because it possesses improved feeding value for poultry (Han Y. et al. (1987) Poultry Sci.
66:103-111) and livestock (Nordstrom, J.W. et al. (1972) J. An. Sci 35(2):357-361). Grain from conventional corn hybrids typically contains 4% oil.
In an effort to increase the kernel oil concentration, a long-term recurrent selection program was initiated in the open-pollinated cv. Burr's White by C.G. Hopkins in 1896. This recurrently-selected population known as Illinois High Oil (IHO), has been selected for increased oil concentration for over ninety generations (Dudley, J.W. and R.J. Lambert. (1992) Maydica 3 7:1-7) using modified mass selection.
As a result, oil concentration was increased in the population over 20%. The germplasm was little used because derived materials had yields substantially lower than conventional varieties (Alexander, D.E. (1988) In: Proc. 43rd Ann.
Corn and Sorghum Res. Conf. Am. Seed Trade Assoc., Washington, D.C.
pp 97-105).
Using thirty-eight open-pollinated cultivars and synthetics, Alexander initiated a second recurrent selection program (Alexho synthetic) to increase kernel oil (Alexander, D.E. (1988) In: Corn and Corn Improvement. G.F. Sprague and J.W. Dudley eds. American Society of Agronomy, Madison WI. Pp 869-880).
Equivalent oil levels to IHO were achieved in twenty-eight cycles using selection based upon the oil concentration of single ears and in later generations based upon the oil concentration of single kernels. Yield performance of Alexho-derived material in single cross hybrids (high oil inbred x conventional inbred) is improved over IHO, presumably due to the greater genetic variability initially available, although performance was not equivalent to conventional hybrids. The development of agronomically elite corn germplasm also containing increased kernel oil concentration is clearly a challenge using conventional plant breeding methods.
Kernel oil concentration can be phenotypically measured using a variety of analytical methods. Oil concentration displays a non-discrete distribution, common for quantitatively-inherited traits controlled by several loci. Kernel oil measurements select those breeding lines with the highest phenotypic expression.
Unfortunately, the genetic potential for high oil is limited in most of these lines because it is impossible to discriminate between lines based upon their true genetic composition. This situation is further aggrevated when simultaneous selection for agronomic performance is practiced. It would therefore be advantageous to base selection upon the genotype of the plants in the population.
Genetic markers, especially nucleic acid markers, may be used to advantage as an indirect selection method for complex quantitative traits. Genetic markers identifying alleles conferring increased oil would therefore be an advantageous tool for plant breeding programs developing elite high oil corn germplasm.
There is limited published information on the identification of genetic markers predictive for increased oil yield. Kahler (Kahler, A.L. (1985) In: Proc.
Ann. Corn and Sorghum Res. Conf. Am. Seed Trade Assoc., Washington D.C. pp. 66-89) measured isozyme allelic frequency changes following twentyfive cycles of selection in Alexho synthetic and found eight significant loci. Most of these allele frequency changes were also significant for tests measuring random genetic drift, making it difficult to conclude that selection based upon these isozyme alleles would be useful. More recently Goldman et al. (Goldman. et al. (1994) Crop Sci. 34:908-915) and Berke and Rocheford (Berke, T.G. and Rocheford, T.R. (1995) Crop Sci. 35:1542-1549) used RFLP markers to identify S: significant marker loci associated with oil concentration in the Illinois long-term selection populations. These studies identified twenty-five and thirty-one markers respectively, in populations derived from Burr's White, which were significantly '20 associated with increased oil. Some of the regions identified by significant RFLP :marker loci may be in common between the two studies, however of the fifteen RFLP markers which were used in both studies, six were in disagreement for their effect on oil concentration. In these studies the populations used were derived from common ancestry (Burr's White); however, the populations were selected for 25 different traits (oil and protein) over many generations. It is not surprising that many identified oil loci would be unique to each population analyzed. It is therefore desirable to identify those genetic markers which are uniquely predictive of germplasm being used in the breeding program.
SUMMARY OF INVENTION A method is disclosed for reliably and predictably breeding for corn with increased kernel oil concentration. The method comprises a) using a pair of primers for one or more genetic markers to select a corn plant from a corn breeding population by marker-assisted selection, wherein said primers are selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 15 and 16, SEQ ID ck10764apr4.speci NOS: 17 and 18, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 and 28, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 and 34, SEQ ID NOS: 35 and 36, SEQ ID NOS: 37 and 38, SEQ ID NOS: 39 and 40, SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and 44, SEQ ID NOS: 45 and 46, SEQ ID NOS: 47 and 48, SEQ ID NOS: 49 and 50, SEQ ID NOS: 51 and 52, SEQ ID NOS: 53 and 54, SEQ ID NOS: 55 and 56, SEQ ID NOS: 57 and 58, SEQ ID NOS: 59 and SEQ ID NOS: 61 and 62, SEQ ID NOS: 63 and 64, SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and 68, SEQ ID NOS: 69 and 70, SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and 74, SEQ ID NOS: 75 and 76, SEQ ID NOS: 77 and 78, SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104; and b) crossing the selected corn plant with a second corn plant wherein the progeny of the cross displays increased kernel oil concentration. A preferred source of 1' high oil corn germplasm is a member of an Alexho synthetic population or a progeny thereof Also disclosed is a method for identifying corn plants or corn lines for use as parents for creation of a breeding population, the method comprising a) genotyping corn plants or corn lines with one or more genetic markers wherein the genetic markers are selected from the S group consisting ofsl375, s1384, s1394, s1416, s1422, s1432, s1457, s1480, s1476, s1478, 20 s1484, sl500, s1513, s1529, s1544, s1545, s1630, s1633, s1647, s1750, s1756, s1757, s1767, s1772, s1774, s1780, s1797, s1813, s1816, s1817 s1836, s1853, s1860, s1870, s1921, s1922, s1925, s1931, s1933, s1939, s1946, s1949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175, using a pair of primers for each said genetic marker, said primers selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 15 and 16, SEQ ID NOS: 17 and 18, SEQ ID NOS: 19 and SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 and 28, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 and 34, SEQ ID NOS: 35 and 36, SEQ ID NOS: 37 and 38, SEQ ID NOS: 39 and 40, SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and 44, SEQ ID NOS: 45 and 46, SEQ ID NOS: 47 and 48, SEQ -3A- ID NOS: 49 and 50, SEQ ID NOS: 51 and 52, SEQ ID NOS: 53 and 54, SEQ ID NOS: 55 and 56, SEQ ID NOS: 57 and 58, SEQ ID NOS: 59 and 60, SEQ ID NOS: 61 and 62, SEQ ID NOS: 63 and 64, SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and 68, SEQ ID NOS: 69 and SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and 74, SEQ ID NOS: 75 and 76, SEQ ID NOS: 77 and 78, SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104, respectively; and b) identifying corn plants or corn lines which, based upon their genotype, are predicted to produce transgressive segregants for kernel oil concentration.
The present invention provides a method for the identification of and selection for genes controlling increased corn kernel oil concentration. These oil alleles were initially identified in materials composed of or derived from the Alexho synthetic breeding 1 populations. Further, the method facilitates the use of this high oil material in breeding programs with the objective of developing new high oil corn germplasm.
SSpecifically, the method uses genetic markers to predict the oil breeding value of lines in a corn breeding program. By indirect selection of oil loci using these markers, those lines with the greatest genetic potential for increased kernel oil concentration are chosen.
According to the method, any type of genetic marker may be used to identify an 20 association with kernel oil concentration. The method is only limited by the ability to measure S polymorphism at a given marker locus. Those skilled in the art will recognise that the various genetic markers which may be used includes but is not limited to restriction fragment length polymorphisms (RFLPs), random amplified polymorphic DNAs (RAPDs), simple sequence repeats (SSRs), AFLPs, various single base pair detection methods, allozymes, and phenotypic markers. SSR markers useful in the practice of the instant method include s1375, s1384, s1394, s1416, s1422, s1432, s1457, s1476, s1478, s1480, s1484, s1500, s1513, s1529, s1544, s1545, s1630, s1633, s647, s750, s756, s757,767, s772, s1774, s1780, s1797, s1813, s1816, s1817, s1836, s1853, s1860, s1870, s1921, s1922, s1925, s1931, s1933, s1939, s1946, s1949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and 2175.
WO 98/42870 PCT/US98/05550 A further embodiment of the present invention are the trait loci controlling the expression of corn kernel oil concentration. These loci are identified and defined mapped) by the marker loci of the present invention.
An additional embodiment of the present invention are corn plants and high oil corn germplasm that are produced using the instant breeding method.
DETAILED DESCRIPTION OF THE INVENTION Table 1 provides a brief description of the genetic markers that form a part of the instant invention. Each marker is defined by it's constituent nucleic acid primers (forward and reverse) that facilitate amplification of the specific marker locus of the corn genome. Also indicated is the required identifier for each sequence. The identifiers listed in Table 1 correspond to those listed in the Sequence Listing (infra) as required by 37 C.F.R. §1.821 et seq.
Table 1 Genetic markers useful for defining the location of trait loci controlling corn kernel oil concentration Marker Sequence Primer Type SEQ ID NO.
s1375 TTTATGGGTTGGGAGATACTTG forward 1 AGATGTGTGCGTTTTTGAGAG reverse 2 s1384 TTACGGCCTAGACATTTCGAC forward 3 CACTTGCTTTCAGGTACCCA reverse 4 s1394 CTGCCCAGTCCGTAATGAA forward TAGATTATTTTCTGAACGATTGG reverse 6 s1416 GATCTCTCTGAGGCTTGTCC forward 7 TGTAGTTGAGGATGCTCCC reverse 8 s1422 AGGCAAGGCTTTCTTCATAC forward 9 CGGACGACGACTGTGTTC reverse s1432 ACATGAGAAACAAGATAGAACCAG forward 11 AAAATGTAAGAACTTGTTTGGGA reverse 12 s1457 CTGCTTATTGCTTTCGTCATA forward 13 TGCTGCACTACTTGAACCTAG reverse 14 s1476 ACACAGAGATGACAAAAGCAA forward GCAGGCGTGCTATGAGAG reverse 16 s1478 AGCGGTGAAACCCTTATG forward 17 CTGTGGCTGGTTCCTCTC reverse 18 s1480 GCTCTTGATAAAAAGGCAAGT forward 19 CTTGTTGTAATGGATGAGTGAG reverse s1484 GCTCGTAGTAGGGGTTACG forward 21 GACAGCCTCACCTCAAGA reverse 22 WO 98/42870 PCT/US98/05550 ACAGATCTTGACACGTACATACC forward 23 GGACGTGTATCCTCAAATCAT -~-reverse 24 s1513 CAGCGAATACTGAATAACGC forward TGTTGGATGAGCACTGAAC reverse 26 s1529 TGTTCTCAACAACCACCG forward 27 CGTTTAGCGATATCATTTCC reverse 28 s1544 GATCCTACCAAATCTTATAGGC forward 29 ACAGCTAGCCAAGATCTGATTl reverse s1545 CGATACTAATGGAAGCCCTA forward 31 ATGGCCCAT'TAAGTTTATCAC reverse 32 s1630 AAAGCGTAGTCGGpA&AGC forward 33 ACCAATGATCT-TACGCAGAT reverse 34 s1633 TAATCAGAGCGTACATCAGGA forward AGGGCATCAATCAAGATG reverse 36 s1647 GAGAC=IrGAGGAGAAAGCA forward. 37 GATCAAAAGAGCAAAAGGAGA reverse 38 s1750 AACTGATGAATACCTTCCCAG forward 39 TGATTAACTTfCTCCCTTTGGT reverse s1756 TGGGCACAACATATGAGT-rAC forward 41 CCCCCATAGAGAGAGATAGAG reverse 42 s1757 AAGCACGGCCCAATAGAAT forward 43 AGGATGTCCCTAGCTfl'Ayj' reverse 44 s1767 TCATTGCCCAAAGTGTTG forward CTCATCACCCGTGCAGAG reverse 46 s1772 GATCCACGCCATTTAAAC forward 47 TGATACTCTGGTGCATGTTC reverse 48 s1774 GATCGCTCCGATCTATCG forward 49 AGCGGCATCTATGTTCTATG reverse s1780 CCCAGTGCGAAGAGACTC forward 51 ACACCTGCTCTGCACCAC reverse 52 s1797 CTAACCCACGACGACCCT forward 53 GCATGAGTGCATGTGCAT reverse 54 s1813 CTGCCACATGCTTTTCTG forward .CTGTAAAGAAGCTGGTCTGGA reverse___*- 56 s1816 TTCTCCTCATGGATGCGT forward 57 CTATTTGGAAGTATGGGCTTCA reverse 58 WO 98/42870 PTU9/55 PCT/US98/05550 si 817 GAGGGCATCTATGTGCAAC forward 59 GCTCAGAAGTTGCGTTTATG reverse s1836 TTCCTTCACGTTTCTCTGTAA forward 61 CACATAAACCTAATGGGGTACA reverse 62 s1853 CCCAAAGGCGATACCTATT forward 63 CCCACTTTCTCACTCTTTCT reverse 64 s1860 GAGGTGAGTACTATGCAAATGC forward CAGGCTTACCTAGCCTTCTC -reverse 66 s1870 CTATGGATGGCTGCTTGC forward 67 GTCAGGCAGCAGAATGTG -_-reverse 68 ___68 s1921 AAACCGTCCAGCCJACTAC forward 69 GGAAGAACCAATCCCATATCT reverse s1922 AACATCCTGTCGGAAACAG forward 71 TCATCACGTCTCTCTTCAAC reverse 72 s1925 TTGTGGCAGAATCTCAAATTA forward 73 CGACTGGTGACATGTCJAAG reverse 74 s1931 AGTGAGGAAAGAATATGCTGG forward TGGACTGAGAAACTGATTTGA reverse 76 s1933 CACAAATGTGAAGGTAAACACT forward 77 AATGGTACGGTTCAGGATG reverse 78 s1939 AGATGACGCACGGAACAC forward 79 AGCATCATGTAGCAGGAGG -reverse s1946 TTGCAGCACTGTCGTAGTC forward 81 GCGCGAGTGGAGTAGTAAG reverse 82 s1949 AAGA'ITATGCAGATGAGACACC forward 83 GTTCCATGCTITICCTTGG reverse 84 s2054 GCCGATACCATGTAAGAGAAT forward CTCTGGGCTCTGTGTTAGAGT reverse 86 s2055 CTGCTTTCTCTGTTCCAGC forward 87 AATCGCTTACTTGTAACCCAC reverse 88 s2057 AAGAACGTACGTCCCATAAAG forward 89 CAAGGTAAAGTGACAAAGCAG reverse s2058 GTTCAGGATGAGGCGGAjA forward 91 GTGATCATCGCAGGAGACC __reverse 92 s2097 GGAGCCTGGAGTGAGAAC forward 93 CATGCTCACCTAACGTGG reverse 94 WO 98/42870 PCT/US98/05550 s2122 ATCTGAACACTTGAGCAACAA forward ATAGACCGGACCCATCAC reverse 96 s2125 CGAACAGCGGGTACACCT forward 97 GAGGTCAGCTTCCTCGATCT reverse 98 s2150 GGAATCGTTCCTCCACAC forward 99 CTTCCTCGGTGTCAGACG reverse 100 s2156 ATGGAAACATCAAAGTGGATT forward 101 TGCTACCCTGATGACCTGAT reverse 102 s2175 ACCACTAGTCTCATATGAAGGG forward 103 GGTAGGTGGGTAGGGGTT reverse 104 For the purposes of this invention, we define the following terms: Corn. Any variety, cultivar, or population of Zea mays L.
Elite. This term characterizes a plant or variety possessing favorable traits, such as, but not limited to high yield, good grain quality, and disease resistance.
This enables its use in commercial production of seed or grain at a profit. The term also characterizes parents giving rise to such plants or varieties.
High Oil Corn Germplasm. This term characterizes corn plants which, when either self-pollinated or used as either the male or the female parent in a variety of outcrossing combinations, produce kernels with increased oil when compared to kernels produced by non-high oil germplasm. Examples of high oil corn germplasm include but are not limited to open-pollinated varieties, hybrids, synthetics, inbred lines, races, and populations or corn plants derived from one of the aforementioned.
Variety or cultivar. These terms refer to a group of similar plants that by structural features and performance can be identified from other varieties or cultivars within the same species.
Line. This term refers to a group of individuals from a common ancestry; a more narrowly defined group than a variety.
Synthetic. This term refers to a genetically heterogeneous collection of plants of known ancestry created by the intermating of any combination of inbreds, hybrids, varieties, populations, races or other synthetics.
Inbred. This term refers to a substantially homozygous individual, variety or line.
Recombinant Inbreds. A population of independently derived lines developed by repeated selfing each generation until complete homozygosity is approached. Each recombinant inbred is derived from a single F2 plant using a breeding method commonly referred to as single seed descent.
WO 98/42870 PCTIUS98/05550 Breeding. The art and science of improving a species of plant or animal through controlled genetic manipulation.
Marker-Assisted Selection. The use of genetic markers to identify and select plants with superior phenotypic potential. Genetic marker(s) determined previously to be associated with a trait locus or trait loci are used to uncover the genotype at trait loci by virtue of linkage between the marker locus and the trait locus. Plants containing desired trait alleles are chosen based upon their genotypes at linked marker loci.
Alexho Synthetic. Recurrently selected, high oil corn germplasm developed by Denton Alexander at the University of Illinois. Alexho synthetic high oil corn germplasm is composed of multiple synthetic populations defined by their cycle of advancement in the recurrent selection breeding program.
Breeding Population. A genetically heterogeneous collection of plants created for the purpose of identifying one or more individuals with desired phenotypic characteristics.
Phenotype. The observed expression of one or more plant characteristics.
Phenotypic Value. A measure of the expected expression of an allele at a trait locus. The phenotypic value of an allele at a trait locus is dependent upon its expressive strength in comparison to alternative alleles. The phenotypic value of an individual, and hence its phenotypic potential, is based upon its total genotypic composition at all loci for a given trait.
Transgressive Segregants. Individuals whose phenotype exceeds the phenotypic variation predicted by the parents.
Genetic Marker. Any morphological, biochemical, or nucleic acid based phenotypic difference which reveals a DNA polymorphism. Examples of genetic markers includes but is not limited to RFLPs, RAPDs, allozymes, SSRs, and AFLPs.
Marker locus. The genetically defined location of DNA polymorphisms as revealed by a genetic marker.
Trait Locus. A genetically defined location for a collection of one or more genes (alleles) which contribute to an observed characteristic.
Genotype. The allelic composition of an individual at genetic loci under study.
Restriction Fragment Length Polymorphism (RFLP). A DNA-based genetic marker in which size differences in restriction endonuclease generated DNA fragments are observed via hybridization (Botstein, D. et al. 1980. Am. J.
Hum. Genet. 32: 314-331.
Random Amplified Polymorphic DNA (RAPD). A DNA amplificationbased genetic marker in which short, sequence arbitrary primers are used and the WO 98/42870 PCT/US98/05550 resulting amplification products are size separated and differences in amplification patterns observed (Williams J.G.K. et al. 1990. Nucleic Acids Res. 18:6531-6535).
Simple Sequence Repeat (SSR). A DNA amplification-based genetic marker in which short stretches of tandemly repeated sequence motifs are amplified and the resulting amplification products are size separated and differences in length of the nucleotide repeat are observed (Tautz D. 1989. Nucleic Acids Res. 112:4127-4138).
AFLP. A DNA amplification-based genetic marker in which restriction endonuclease generated DNA fragments are ligated to short DNA fragments which facilitate the amplification of the restricted DNA fragments (Vos, P. et al.
1995. Nucleic Acids Res. 23:4407-4414). The amplified fragments are size separated and differences in amplification patterns observed.
Allozymes. Enzyme variants which are electrophoretically separated and detected via staining for enzymatic activity (Stuber, C.W. and M.M. Goodman.
1983. USDA Agric. Res. Results, Southern Ser., No. 16).
The present invention relates to the discovery of trait loci controlling kernel oil concentration through the use of genetic markers. In populations in which variation for both kernel oil concentration and genetic marker alleles exist, oil measurements and marker-based genotypes were generated for members of the populations. Using least squares methods, the locations of oil concentration loci were determined in relation to markers genetically linked to these trait loci.
Indirect selection of preferred oil alleles may now be practiced using the information at one or more linked genetic markers. Selected corn plants comprise one or more alleles encoding a high oil phenotype.
It is recognized that several different populations and population types could be used to locate trait loci of interest. Some of the population types include but are not limited to recombinant inbreds, backcrosses, F2's or their selfpollinated or intermated derivatives, and synthetics. Further, it is understood that an alternative to measuring phenotypic and genotypic variation within populations is the measurement of genotypes and phenotypes between populations. In this alternative the second population is a selected derivative of the first population, selection being either on the trait of interest (phenotypic selection) or on specific marker alleles (genotypic selection). It is also recognized by those skilled in the art that alternative statistical approaches may be used to determine a linkage relationship between marker loci and trait loci.
EXAMPLES
The present invention is further defined in the following Examples. It should be understood that these Examples, while indicating preferred embodiments of the invention, are given by way of illustration only. From the WO 98/42870 PCT/US98/05550 above discussion and these Examples, one skilled in the art can ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions.
EXAMPLE 1 LOCATION OF LOCI CONFERRING INCREASED KERNEL OIL CONCENTRATION Population development and trait measurement LH119wx and LH51, two inbred corn lines developed by Holden's Foundation Seed Co., Williamsburg, IA were independently intermated with individual plants from the synthetic population ASKC28wx (deposited at the American Type Culture Collection, Rockville, MD; Accession No. ATCC 75105) (waxy kernels are highly represented in ASKC28 and as such I have designated the ASKC28 as being waxy). The Fl plants were selfed and resulting F2 populations were grown. Individual F2 plants were selfed and derived kernels were advanced using single seed decent through six generations of selfing (S6) to produce recombinant inbred lines. Up to twenty kernels from the S6 generation were grown and selfed producing a family of S7 ears representing each recombinant inbred line. Oil values were determined for each ear within a family using near infrared transmitance (Williams, P.C. (1987) In: Near Infrared Technology in the Agricultural and Food Industries; P.C. Williams and C. Norris, eds. American Association of Cereal Chemists).
Genotypic determination Ten seeds from single ears representing each of one hundred ninety-four (LH 119wx x ASKC28wx) or two hundred and four (LH51 x ASKC28wx) recombinant inbred lines were germinated on moistened filter paper. Root segments were excised from germinated seeds, pooled for each ear and extracted using an automated DNA extraction machine. The instrument uses a modification of the Murray and Thompson CTAB procedure (Murray, M.G. and Thompson, W.F. (1980) Nucl. Acids Res. 8:4321-4325). DNA samples were quantified via fluorescence using YoPro-1TM iodide (Molecular Probes, Inc., Eugene, OR) and diluted to 4 pg/ml.
SSR regions for each DNA sample were analyzed using the following protocol: 1. Ten tl of amplification cocktail (see Table 2) was added to 5 .l ng) of extracted DNA; 2. The DNA fragment flanked by sequences complementary to the primers present in the amplification cocktail was amplified by PCR Patent No. 4,683,202 and U.S. Patent No. 4,683,195) using the following protocol: WO 98/42870 PCTIUS98/05550 1) 45 cycles of 50 sec at 95 0 C, 50 sec at 54°C and 80 sec at 72°C and 2) 1 cycle of 300 sec at 72 0
C;
3. Approximately 8 tl of each sample was loaded onto agarose gels composed of 2% Metaphor (FMC Corp., Rockland, ME), IX TBE, and 0.5 Pg/ml ethidium bromide, and electrophoresed for 2 h at 6.1 V/cm in horizontal electrophoresis units to which IX TBE buffer and 0.5 ug/ml ethidium bromide was added; and 4. DNA bands were visualized by UV fluorescence.
Table 2 Amplification Cocktail Reagent Stock Concentration Final Concentration Buffer* 10X dNTPs 2 mM 0.3 mM Forward Primer 40 pLM 0.45 itM Reverse Primer 40 pM 0.45 pM AmpliTaq PolymeraseTM 5 U/tl 0.05 U/pl 10X Buffer is a pH 9.0 solution composed of 800 mM Tris-OH, 200 mM
(NH
4 2 S0 4 and 25 mM MgC1 2 Localization of oil loci One hundred thirty three polymorphic SSR marker loci were used to genotype the recombinant inbreds from the LH119wx x ASKC28wx cross and one hundred and three polymorphic SSR marker loci were used to genotype the LH51 x ASKC28wx-derived population. In addition, twenty publicly available polymorphic SSR loci with previously established chromosome locations and covering all ten maize chromosomes (available from Research Genetics, Huntsville, AL) were also mapped in both populations.
Genetic linkage and distance between marker loci was determined independently for each population using MAPMAKER 3.0 (Lincoln et al.
(1993) Whitehead Inst. Biomed. Res., Cambridge, MA). This resulted in the establishment of ten linkage groups for each population corresponding to the ten chromosomes of maize. Each linkage group was assigned to a chromosome based upon linkage to the public SSR markers. Twenty-three and ten markers in the LH119wx x ASKC28wx and LH51 x ASKC28wx populations, respectively, were not assigned chromosome positions because genetic linkage could not be clearly established.
Analysis of variance was used to identify marker loci in linkage with trait loci conferring increased oil concentration. Oil concentration was used as a WO 98/42870 PCTIUS98/05550 dependent variable and separate ANOVAs were calculated with SAS Proc GLM (SAS Inst., Cary, NC) using each marker locus as a single independent variable (Edwards, et al. (1987) Genetics 116:113-125). Therefore, for each ANOVA test the mean oil values of marker allele classes were compared. Marker loci were declared significant if p 0.05.
Linkage data for significant marker loci was examined to determine both the number of trait loci present and their probable location. Significant marker loci on the same linkage group are either detecting the same trait locus or alternatively different trait loci. By careful examination of the phenotypic variation explained by each marker locus along the chromosome, a determination of the number trait loci on a linkage group was made. Significant marker loci, on the same linkage group and uninterrupted by non-significant marker loci, were declared to be detecting the same trait locus on the chromosome. If significant marker loci on the same chromosome were interrupted by non-significant marker loci then each significant region was declared to contain a trait locus resulting in multiple trait loci on'the same chromosome.
To confirm the number of trait loci, marker data assigned to linkage groups and oil data were also analyzed with Mapmaker/QTL 1.0 (Lincoln, S.E. et al. (1990) Whitehead Inst. Biomed. Res., Cambridge, MA). Results with Mapmaker/QTL were in agreement with the initial analysis for the number trait loci on each chromosome.
Eleven and twelve loci controlling kernel oil concentration were located in the LH l9wx x ASKC28wx and LH51 x ASKC28wx recombinant inbred populations, respectively. Each oil locus is defined by one or more linked marker loci.
In instances where the same marker loci were used in both populations, alignment of linkage groups is possible. It was found that in most instances both populations localized the same oil loci. By considering common marker loci, a total of seventeen loci controlling kernel oil concentration were found. Each oil locus was assigned an arbitrary letter designation (Table 3).
Table 3 Marker loci genetically linked to and predictive of the location of trait loci conferring increased kernel oil concentration Oil locus Chromosome Marker loci A 1 s1922 B 1 s147 8 s853, s949 C 1 S1860, s1925, s1931, s2150 D 2 s2175 WO 98/42870 PCTfUS98/05550 s1394 s1 4 7 6, sl772, s1816, s2122, s1836 s1939, s1946 s1870 s1529 s2054, s1647, s1500, s1545, s1774, s2097 s1457, s2055, s1757, s2125, s1780, s1375, s1797, s1416, s1432, s1921 s1630, s1422, s2156 s1817, s2057 s1544, s1633, s1384, s1813, s1767, s2058, s1933,s1513, s1484 s1756 s1480 (positive oil allele in LH51) s1750 O 10 P 10 Q N.A.* chromosome location not knc )wn In instances where comparisons could be made, oil loci which were identified in one population were identified at the same location in the second population. In two exceptions, an oil locus was found in one population, but not in the second population. In the first case, the allele with a positive oil effect was found in LH51 and thus it would be unexpected to identify the same locus in the LH119wx x ASKC28wx population. In the second case, it was found that different ASKC28wx-derived marker alleles were segregating in the populations; therefore, each population was measuring the oil effect of a different ASKC28wx allele at the trait locus. The most abundant ASKC28wx oil allele segregating in LHI 9wx x ASKC28wx had a positive oil effect versus the alternative LH 119derived allele, whereas in the LH51 x ASKC28wx population, the abundant ASKC28wx allele had no positive oil effect. With the exception of the oil locus linked to marker s1480, all alleles with positive effects on oil concentration were derived from ASKC28wx.
EXAMPLE 2 MARKER-ASSISTED SELECTION OF BREEDING LINES USING GENETIC MARKERS FOR INCREASE KERNEL OIL CONCENTRATION Genetic marker loci in linkage with oil trait loci are highly predictive of oil concentration and as such may be used as an indirect measurement of kernel oil in a marker-assisted selection program. Accordingly, genotypic information from linked marker loci would facilitate the selection of breeding lines with increased oil concentration. Direct oil measurements cannot differentiate between various WO 98/42870 PCT/US98/05550 genotypic trait locus compositions with equivalent phenotypic effects. This is especially problematic in early generation segregating breeding populations where only limited fixation of oil loci has occurred.
By way of example, an objective of a corn breeding program could be the creation of new elite inbred lines which contain trait alleles conferring increased kernel oil concentration. These trait alleles would be introduced by the intermating of high oil germplasm with one or more elite corn inbreds. The resultant hybrid could be self-pollinated to produce an F2 population for the purposes of initiating a conventional pedigree breeding program (Allard, R.W.
(1960) Principles of Plant Breeding. John Wiley Sons, Inc. New York.
Pp 115-128).
In order to identify those F2 individuals with the desired genotypes, plant tissue would be collected from each F2 individual in the population and genotyped with the SSR marker loci listed in Table 1. Those F2 individuals with the highest frequency of SSR marker alleles derived from the high oil source would be selected and further culled based upon their agronomic fitness. With continued inbreeding and segregation, those oil loci in a heterozygous state could become fixed for either the high oil or low oil allele. It is therefore likely that genotyping and selection of later generation materials would be practiced in order to further segregate breeding lines based upon their marker allele and hence oil allele composition.
Depending upon population size and serendipity, the resulting inbreds from the pedigree breeding program may not demonstrate sufficient agronomic competitiveness or sufficient kernel oil expression because an inadequate number of oil alleles was recovered. These new inbreds could therefore be used as parental material and new breeding projects initiated. The SSR markers could again be used for further selection of oil as described.
It is obvious to those skilled in the art that many variants to selection methodology may by envisioned. Selection would be based upon the allelic composition of one or more marker loci which identify trait oil loci present in a population. Further selection would be performed by examination and selection of genotypes from individual plants, families, or their progeny. Various predictive models could be developed using genotypic information, which could generate various selection indices. These models would permit weighting the effect predicted by marker loci. This is because the predictive value of an individual marker locus is dependent upon its genetic distance from the corresponding trait locus as well as the expressivity of the trait locus. Selection strategies which combine phenotype-based and genotype-based selection may also be envisioned.
WO 98/42870 PCTIUS98/05550 The marker loci presented here are predictive of oil loci in Alexho synthetic populations. Because ASKC28wx represents the 28th oil breeding cycle of a genetically closed population, earlier breeding cycles are composed of the same oil loci. It is expected that cycles differ simply in their allelic frequency at the identified oil loci. Therefore, in breeding populations derived from earlier Alexho cycles, the marker loci described in this invention will be useful in identification of oil loci and in prediction of oil concentration.
EXAMPLE 3 IDENTIFICATION OF CORN PLANTS FOR USE AS PARENTS FOR THE PRODUCTION OF TRANSGRESSIVE SEGREGANTS
FOR
KERNEL OIL CONCENTRATION It is important to identify corn plants and lines which, when used as parents, have the greatest probability of producing offspring with superior performance. Transgressive segregant offspring of such parents would result from the crossing of parents with complementary sets of alleles conferring the high-oil phenotype. Using the information provided herein, marker alleles which predict desired trait performance high oil) at a given marker locus are known. By genotyping lines at those marker loci, the value of those lines as parents is revealed. For example, if one wanted to create an individual containing superior alleles at 5 separate oil loci one could identify and cross a parent composed of desired alleles for locus A, B, and C with a parent composed of desired alleles at B, D, and E. These parents are complementary because they permit the recovery of progeny containing desired alleles at all 5 loci. Ideally, parents would be chosen which when combined ensure maximum complementation of loci, so that a high frequency of desired recombinants are recovered.
WO 98/42870 PCT[US98/05550 SEQUENCE LISTING GENERAL INFORMATION:
APPLICANT:
ADDRESSEE: E. I. DU PONT DE STREET: 1007 MARKET STREET CITY: WILMINGTON STATE: DELAWARE COUNTRY: USA ZIP: 19898 TELEPHONE: 302-992-4926 TELEFAX: 302-773-0164 TELEX: 6717325 NEMOURS AND COMPANY (ii) TITLE OF INVENTION: A METHOD TO IDENTIFY AND BREED CORN WITH INCREASED KERNEL OIL
CONCENTRATION
(iii) NUMBER OF SEQUENCES: 104 (iv) COMPUTER READABLE FORM: MEDIUM TYPE DISKETTE, 3.50 INCH COMPUTER: IBM PC COMPATIBLE OPERATING SYSTEM: MICROSOFT WINDOWS SOFTWARE: MICROSOFT WORD VERSION 7.OA CURRENT APPLICATION DATA: APPLICATION NUMBER: FILING DATE:
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CLASSIFICATION:
60/041,515 24, 1997 (vii) ATTORNEY/AGENT INFORMATION: NAME: MAJARIAN, WILLIAM R.
REGISTRATION NUMBER: P-41,173 REFERENCE/DOCKET NUMBER: BB-1076 WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID NO:1: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1: TTTATGGGTT GGGAGATACT TG 22 INFORMATION FOR SEQ ID NO:2: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2: AGATGTGTGC GTTTTTGAGA G 21 INFORMATION FOR SEQ ID NO:3: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3: TTACGGCCTA GACATTTCGA C 21 INFORMATION FOR SEQ ID NO:4: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: CACTTGCTTT CAGGTACCCA WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CTGCCCAGTC CGTAATGAA 19 INFORMATION FOR SEQ ID NO:6: SEQUENCE CHARACTERISTICS: LENGTH: 24 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6: TAGATTTATT TTCTGAACGA TTGG 24 INFORMATION FOR SEQ ID NO:7: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7: GATCTCTCTG AGGCTTGTCC INFORMATION FOR SEQ ID NO:8: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8: TGTAGTTGAG GATGCTCCC. 19 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:9: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9: AGGCAAGGCT TTCTTCATAC INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other.nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CGGACGACGA CTGTGTTC 18 INFORMATION FOR SEQ ID NO:11: SEQUENCE CHARACTERISTICS: LENGTH: 24 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11: ACATGAGAAA CAAGATAGAA CCAG 24 INFORMATION FOR SEQ ID NO:12: SEQUENCE CHARACTERISTICS: LENGTH: 23 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12: AAAATGTAAG AACTTGTTTG GGA 23 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:13: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13: CTGCTTATTG CTTTCGTCAT A 21 INFORMATION FOR SEQ ID NO:14: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14: TGCTGCACTA CTTGAACCTA G 21 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID ACACAGAGAT GACAAAAGCA A 21 INFORMATION FOR SEQ ID NO:16: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16: GCAGGCGTGC TATGAGAG 18 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:17: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17: AGCGGTGAAA CCCTTATG 18 INFORMATION FOR SEQ ID NO:18: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18: CTGTGGCTGG TTCCTCTC 18 INFORMATION FOR SEQ ID NO:19: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19: GCTCTTGATA AAAAGGCAAG T 21 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE:. other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CTTGTTGTAA TGGATGAGTG AG 22 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:21: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21: GCTCGTAGTA GGGGTTACG 19 INFORMATION FOR SEQ ID.NO:22: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22: GACAGCCTCA CCTCAAGA 18 INFORMATION FOR SEQ ID NO:23: SEQUENCE CHARACTERISTICS: LENGTH: 23 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23: ACAGATCTTG ACACGTACAT ACC 23 INFORMATION FOR SEQ ID NO:24: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24: GGACGTGTAT CCTCAAATCA T 21 WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CAGCGAATAC TGAATAACGC INFORMATION FOR SEQ ID NO:26: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26: TGTTGGATGA GCACTGAAC INFORMATION FOR SEQ ID NO:27: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27: TGTTCTCAAC AACCACCG INFORMATION FOR SEQ ID NO:28: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28: CGTTTAGCGA TATCATTTTC C WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID NO:29: SEQUENCE CHARACTERISTICS: LENGTH: 23 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29: GATCCTACCA AAATCTTATA GGC 23 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID ACAGCTAGCC AAGATCTGAT T 21 INFORMATION FOR SEQ ID NO:31: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31: CGATACTAAT GGAAGCCCTA A 21 INFORMATION FOR SEQ ID NO:32: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE:- other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32: ATGGCCCATT AAGTTTATCA C 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:33: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33: AAAGCGTAGT CGGAAAGC 18 INFORMATION FOR SEQ ID NO:34: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:34: ACCAATGATC TTTACGCAGA T 21 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID TAATCAGAGC GTACATCAGG A 21 INFORMATION FOR SEQ ID NO:36: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:36: AGGGCATCAA TCAAGAATG 19 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:37: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:37: GAGACTTTTG AGGAGAAAGC A 21 INFORMATION FOR SEQ ID NO:38: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:38: GATCAAAAGA GCAAAAGGAG A 21 INFORMATION FOR SEQ ID NO:39: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:39: AACTGATGAA TACCTTCCCA G 21 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID TGATTAACTT CTCCCTTTGG T 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:41: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:41: TCGGCACAAC ATATGAGTTA C 21 INFORMATION FOR SEQ ID NO:42: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:42: CCCCCATAGA GAGAGATAGA G 21 INFORMATION FOR SEQ ID NO:43: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:43: AAGCACGGCC CAATAGAAT 19 INFORMATION FOR SEQ ID NO:44: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:44: AGGATGTCCC TAGCTTTATT G 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID TCATTGCCCA AAGTGTTG 18 INFORMATION FOR SEQ ID NO:46: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:46: CTCATCACCC CTCCAGAG 18 INFORMATION FOR SEQ ID NO:47: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:47: GATCCACGCC ATTTAAAC 18 INFORMATION FOR SEQ ID NO:48: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:48: TGATACTCTG GTGCATGTTC WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:49: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:49: GATCGCTCCG ATCTATCC 18 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID AGCGGCATCT ATGTTCTATG INFORMATION FOR SEQ ID NO:51: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS; single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:51: CCCAGTGCGA AGAGACTC 18 INFORMATION FOR SEQ ID NO:52: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:52: ACACCTGCTC TGCACCAC 18 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:53: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:53: CTAACCCACG ACGACCCT 18 INFORMATION FOR SEQ ID NO:54: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:54: GCATGAGTGC ATGTGCAT 18 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CTGCCACATG CTTTTCTG 18 INFORMATION FOR SEQ ID NO:56: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:56: CTGTAAAGAA GCTGGTCTGG A 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:57: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:57: TTCTCCTCAT GGATGCGT 18 INFORMATION FOR SEQ ID NO:58: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:58: CTATTTGGAA GTATGGGCTT CA 22 INFORMATION FOR SEQ ID NO:59: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:59: GAGGGCATCT ATGTGCAAC 19 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID GCTCAGAAGT TGCGTTTATG WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:61: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:61: TTCCTTCACG TTTCTCTGTT AA 22 INFORMATION FOR SEQ ID NO:62: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:62: CACATAAACC TAATGGGGTA CA 22 INFORMATION FOR SEQ ID NO:63: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS:• single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:63: CCCAAAGGCG ATACCTATT 19 INFORMATION FOR SEQ ID NO:64: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:64: CCCACTTTCT CACTCTTTTC T 21 WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID GAGGTGAGTA CTATGCAAAT GC INFORMATION FOR SEQ ID NO:66: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:66: CAGGCTTACC TAGCCTTCTC INFORMATION FOR SEQ ID NO:67: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:67: CTATGGATGG CTGCTTGC INFORMATION FOR SEQ ID NO:68: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:68: GTCAGGCAGC AGAATGTG WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID NO:69: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:69: AAACCGTCCA GCGACTAC INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID GGAAGAACCA ATCCCATATC
T
INFORMATION FOR SEQ ID NO:71: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:71: AACATCCTGT CGGAAACAG INFORMATION FOR SEQ ID NO:72: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:72: TCATCACGTC TCTCTTTCAA C WO 98/42870 PCTIS98/05550 INFORMATION FOR SEQ ID NO:73: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:73: TTGTGGCAGA ATCTCAAATT A 21 INFORMATION FOR SEQ ID NO:74: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:74: CGACTGGTGA CATGTGAAG 19 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID AGTGAGGAAA GAATATGCTG G 21 INFORMATION FOR SEQ ID NO:76: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:76: TGGACTGAGA AACTGATTTG A 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:77: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:77: CACAAATGTG AAGGTAAACA CT 22 INFORMATION FOR SEQ ID NO:78: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:78: AATGGTACGG TTCAGGATG 19 INFORMATION FOR SEQ ID NO:79: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:79: AGATGACGCA CGGAACAC 18 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID AGCATCATGT AGCAGGAGG 19 WO 98/42870 PCTIUS98/05550 INFORMATION FOR SEQ ID NO:81: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:81: TTGCAGCACT GTCGTAGTC 19 INFORMATION FOR SEQ ID NO:82: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:82: GCGCGAGTGG AGTAGTAAG 19 INFORMATION FOR SEQ ID NO:83: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:83: AAGATTATGC AGATGAGACA CC 22 INFORMATION FOR SEQ ID NO:84: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:84: GTTCCATGCT TTCCTTGG 18 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CTCTGGGCTC TGTGTTAGAG T 21 INFORMATION FOR SEQ ID NO:86: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:86: CTCTGGGCTC TGTGTTAGAG T 21 INFORMATION FOR SEQ ID NO:87: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:87: CTGCTTTCTC TGTTCCAGC 19 INFORMATION FOR SEQ ID NO:88: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE:* other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:88: AATCGCTTAC TTGTAACCCA C 21 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:89: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:89: AAGAACGTAC GTCCCATAAA G 21 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID CAAGGTAAAG TGACAAAGCA G 21 INFORMATION FOR SEQ ID NO:91: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:91: GTTCAGGATG AGGCGGAA 18 INFORMATION FOR SEQ ID NO:92: SEQUENCE CHARACTERISTICS: LENGTH: 19 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:92: GTGATCATCG CAGGAGACC 19 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:93: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:93: GGAGCCTGGA GTGAGAAC 18 INFORMATION FOR SEQ ID NO:94: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:94: CATGCTCACC TAACGTGG 18 INFORMATION FOR SEQ ID SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID ATCTGAACAC TTGAGCAACA A 21 INFORMATION FOR SEQ ID NO:96: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:96: ATAGACCGGA CCCATCAC 18 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:97: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:97: CGAACAGCGG GTACACCT 18 INFORMATION FOR SEQ ID NO:98: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:98: GAGGTCAGCT TCCTCGATCT INFORMATION FOR SEQ ID NO:99: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:99: GGAATCGTTC CTCCACAC 18 INFORMATION FOR SEQ ID NO:100: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:100: CTTCCTCGGT GTCAGACG 18 WO 98/42870 PCT/US98/05550 INFORMATION FOR SEQ ID NO:101: SEQUENCE CHARACTERISTICS: LENGTH: 21 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:101: ATGGAAACAT CAAAGTGGAT T 21 INFORMATION FOR SEQ ID NO:102: SEQUENCE CHARACTERISTICS: LENGTH: 20 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:102: TGCTACCCTG ATGACCTGAT INFORMATION FOR SEQ ID NO:103: SEQUENCE CHARACTERISTICS: LENGTH: 22 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:103: ACCACTAGTC TCATATGAAG GG 22 INFORMATION FOR SEQ ID NO:104: SEQUENCE CHARACTERISTICS: LENGTH: 18 base pairs TYPE: nucleic acid STRANDEDNESS: single TOPOLOGY: linear (ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:104: GGTAGGTGGG TAGGGGTT 18 Where the terms "comprise", "comprises", "comprised" or "comprising" are used in this specification, they are to be interpreted as specifying the presence of the stated features, integers, steps or components referred to, but not to preclude the presence or addition of one or more other feature, integer, step, component or group thereof.
S*
1 -43a-

Claims (25)

1. A method of breeding corn with increased kernel oil concentration comprising: a) using a pair of primers for one or more genetic markers to select a corn plant from a corn breeding population by marker-assisted selection, wherein said primers are selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 5 *25 and 16, SEQ ID NOS: 17 and 18, and 22, SEQ ID NOS: 23 and 24, and 28, SEQ ID NOS: 29 and 30, and 34, SEQ ID NOS: 35 and 36, and 40, SEQ ID NOS: 41 and 42, and 46, SEQ ID NOS: 47 and 48, and 52, SEQ ID NOS: 53 and 54, and 58, SEQ ID NOS: 59 and 60, and 64, SEQ ID NOS: 65 and 66, and 70, SEQ ID NOS: 71 and 72, and 76, SEQ ID NOS: 77 and 78, and 82, SEQ ID NOS: 83 and 84, and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 SEQ ID NOS: 37 and 38, SEQ ID NOS: 39 SEQ ID NOS: 43 and 44, SEQ ID NOS: SEQ ID NOS: 49 and 50, SEQ ID NOS: 51 SEQ ID NOS: 55 and 56, SEQ ID NOS: 57 SEQ ID NOS: 61 and 62, SEQ ID NOS: 63 SEQ ID NOS: 67 and 68, SEQ ID NOS: 69 SEQ ID NOS: 73 and 74, SEQ ID NOS: SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104; and b) crossing the selected corn plant with a second corn plant wherein the progeny corn plants of the cross display increased kernel oil concentration.
2. The method of claim 1 wherein the selected corn plant is member of an Alexho synthetic population or a progeny thereof.
3. A method for identifying corn plants or corn lines for use as parents for creation of a breeding population, the method comprising: a) genotyping corn plants or corn lines with one or more genetic markers wherein the genetic markers are selected from the group consisting of s1375, sl384, 45 s1394, s1416, s1422, s1432, s1457, s1480,.s1476, s1478, s1484, s15QO, s15 13, s1529, s1544, s1545, s1630, s1633, s1647, s1750, s1756, s1757, s1767, s1772, s1774, s1780, s1797, s1813, s1816, s1817 s1836, s1853, s1860, s1870, s1921, s1922, s1925, s1931, s1933, s1939, s1946, s1949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175, using a pair of primers for each said genetic marker, said primers selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ED NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ED NOS: I11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 15 and 16, SEQ ID NOS: 17 and 18, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 and 28, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 and 34, SEQ 110 NOS: 35 and 36, SEQ ID) NOS: 37 and 38, SEQ ID NOS: 39 and 40, SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and 44, SEQ ED NOS: 45 and 46, SEQ ID) NOS: 47 and 48, SEQ ID NOS: 49 and SEQ ID) NOS: 51 and 52, SEQ ID NOS: 53 and 54, SEQ ID) NOS: 55 and 56, SE IDNS 7 n 8 SEQ ID NOS: 59i and 60, SEQ ED NOS: 61 and 62, SEQ ID NOS: 63 and 64, SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and 68 SE DNS.9ad7,.QI OS 1ad7,SQI O:7 n 68, SEQ ED NOS: 69 and 70, SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and 80 7, SEQ ID NOS: 75 and 76, SEQ ID NOS: 77 and 84, SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID) NOS: 95 and 90, SEQ ID NOS: 91 and 98, SEQ ID NOS: 99 and 100, SEQ ED NOS: 101 and 102, SEQ lID NOS: 103 and 104, respectively; and b) identifying corn plants or corn lines which, based upon their genotype, are predicted to produce transgressive segregants for kernel oil concentration.
4. A corn plant produced by the method of claim I having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s2054, s1647, s1500, s1545, s1774 and s2097. -,46- A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s1817 and s2057.
6. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1860, s1931, s2150 and s1925.
7. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1457, s2055, s1757, s2125, s1780, s1375, s1797, s1416, s1432 and s1921.
8. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1544, s1633, s1384, s1813, s1767, s2058, s1933, s1513 and s1484.
9. A corn plant produced by the method of claim 1 having a trait locus controlling kernel :i 5 oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1476, s1772, s1816, s2122 and s1836.
10. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1939 and s1946.
11. A corn plant produced by the method of claim 1 having a trait locus controlling kernel S• oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1478, s1853 and s1949.
12. A corn plant produced by the method of claim 1 having a trait locus controlling kernel g. "oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1630, s1422 and s2156.
13. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1756.
14. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1922.
15. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1529. -47-
16. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1394.
17. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1750.
18. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1870.
19. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s2175. Corn plants that display increased kernel oil concentration produced by the method of claim 1.
21. A method of breeding corn with increased kernel oil concentration comprising: a) using a pair of primers for one or more genetic markers to select a corn plant from a corn breeding population by marker-assisted selection, wherein said primers are selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: and 16, SEQ ID NOS: 17 and 18, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 and 28, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 and 34, SEQ ID NOS: 35 and 36, SEQ ID NOS: 37 and 38, SEQ ID NOS: 39 and 40, SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and 44, SEQ ID NOS: and 46, SEQ ID NOS: 47 and 48, SEQ D NOS: 49 and 50, SEQ D NOS: 51 and 52, SEQ ID NOS: 53 and 54, SEQ ID NOS: 55 and 56, SEQ ID NOS: 57 and 58, SEQ ID NOS: 59 and 60, SEQ ID NOS: 61 and 62, SEQ ID NOS: 63 and 64, SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and 68, SEQ ID NOS: 69 *and 70, SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and 74, SEQ ID NOS: and 76, SEQ ID NOS: 77 and 78, SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104; SEQ ID NOS: 9 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104; and -48- b) crossing the selected corn plant with a second corn plant wherein the progeny corn plants of the cross display increased kernel oil concentration substantially as herein described with reference to the accompanying Examples.
22. A method of identifying corn plants or corn lines for use as parents for creation of a breeding population, the method comprising: a) genotyping corn plants or corn lines with one or more genetic markers wherein the genetic markers are selected from the group consisting of s1375, s1384, s1394, s1416, s1422, s1432, s1457, s1480, s1476, s1478, s1484, s1500, s1513, s1529, s1544, s1545, s1630, s1633, s1647, s1750, s1756, s1757, s1767, s1772, s1774, s1780, s1797, s1813, s1816, s1817 s1836, s1853, s1860, s1870, s1921, s1922, s1925, s1931, s1933, s1939, s1946, s1949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175, using a pair of primers for each said genetic marker, said primers selected from the group consisting of SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 15 and 16, SEQ ID NOS: 17 and 18, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and e S *SS* S *SS* 26, SEQ ID NOS: 27 and 28, 32, SEQ ID NOS: 33 and 34, 38, SEQ ID NOS: 39 and 40, 44, SEQ ID NOS: 45 and 46, 50, SEQ ID NOS: 51 and 52, 56, SEQ ID NOS: 57 and 58, 62, SEQ ID NOS: 63 and 64, 68, SEQ ID NOS: 69 and 70, 74, SEQ ID NOS: 75 and 76, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and SEQ ID NOS: 35 and 36, SEQ ID NOS: 37 and SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and SEQ ID NOS: 47 and 48, SEQ ID NOS: 49 and SEQ ID NOS: 53 and 54, SEQ ID NOS: 55 and SEQ ID NOS: 59 and 60, SEQ ID NOS: 61 and SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and SEQ ID NOS: 77 and 78, SEQ ID NOS: 79 and SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and 104, respectively; and -49- b) identifying corn plants or corn lines which, based upon their genotype, are predicted to produce transgressive segregants for kernel oil concentration substantially as herein described with reference to the accompanying Examples.
23. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s2054, s1647, s1500, s1545, s1774 and s2097 substantially as herein described with reference to the accompanying Examples.
24. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofsl817 and s2057 substantially as herein described with reference to the accompanying Examples. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s1860, s193 1, s2150 and s1925 substantially as herein described with "reference to the accompanying Examples. .i 26. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group S"consisting ofs1457, s2055, s1757, s2125, s1780, s1375, s1797, s1416, s1432 and s1921 substantially as herein described with reference to the accompanying Examples.
27. A corn plant produced by the method of claim 1 having a trait locus controlling kernel S•oil concentration, the locus mapped by a genetic marker selected from the group
99.9 consisting ofs1544, s1633, s1384, s1813, s1767, s2058, s1933, s1513 and s1484 substantially as herein described with reference to the accompanying Examples. 28. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s1476, s1772, s 1816, s2122 and s1836 substantially as herein described with reference to the accompanying Examples. 29. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s1939 and s1946 substantially as herein described with reference to the accompanying Examples. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting ofs1478, s1853 and s1949 substantially as herein described with reference to the accompanying Examples. 31. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s1630, s1422 and s2156 substantially as herein described with reference to the accompanying Examples. 32. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1756 substantially as herein described with reference to the accompanying Examples. 33. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1922 substantially as herein described with reference to the accompanying Examples. 34. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1529 substantially as herein described with reference to the accompanying Examples. 35. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1394 substantially as herein described with reference to the accompanying Examples. 36. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s1750 substantially as herein described with reference to the accompanying Examples. 37. A corn plant produced by the method of claim 1 having a trait locus controlling kernel 25 oil concentration, the locus mapped by the genetic marker s1870 substantially as herein described with reference to the accompanying Examples. 38. A corn plant produced by the method of claim 1 having a trait locus controlling kernel oil concentration, the locus mapped by the genetic marker s2175 substantially as herein described with reference to the accompanying Examples. -51 39. Corn plants that display increased kernel oil concentration produced by the method of claim 1 substantially as herein described with reference to the accompanying Examples. A trait locus controlling kernel oil concentration the locus mapped by a genetic marker defined by a pair of primers selected from SEQ ID NOS: 1 and 2, SEQ ID NOS: 3 and 4, SEQ ID NOS: 5 and 6, SEQ ID NOS: 7 and 8, SEQ ID NOS: 9 and 10, SEQ ID NOS: 11 and 12, SEQ ID NOS: 13 and 14, SEQ ID NOS: 15 and 16, SEQ ID NOS: 17 and 18, SEQ ID NOS: 19 and 20, SEQ ID NOS: 21 and 22, SEQ ID NOS: 23 and 24, SEQ ID NOS: 25 and 26, SEQ ID NOS: 27 and 28, SEQ ID NOS: 29 and 30, SEQ ID NOS: 31 and 32, SEQ ID NOS: 33 and 34, SEQ ID NOS: 35 and 36, SEQ ID NOS: 37 and 38, SEQ ID NOS: 39 and 40, SEQ ID NOS: 41 and 42, SEQ ID NOS: 43 and 44, SEQ ID NOS: 45 and 46, SEQ ID NOS: 47 and 48, SEQ ID NOS: 49 and 50, SEQ ID NOS: 51 and 52, SEQ ID NOS: 53 and 54, SEQ ID NOS: 55 and 56, SEQ ID NOS: 57 and 58, SEQ ID NOS: 59 and 60, SEQ ID NOS: 61 and 62, SEQ ID NOS: 63 and 64, J5 SEQ ID NOS: 65 and 66, SEQ ID NOS: 67 and 68, SEQ ID NOS: 69 and 70, SEQ ID NOS: 71 and 72, SEQ ID NOS: 73 and 74, SEQ ID NOS: 75 and 76, SEQ ID NOS: 77 and 78, SEQ ID NOS: 79 and 80, SEQ ID NOS: 81 and 82, SEQ ID NOS: 83 and 84, SEQ ID NOS: 85 and 86, SEQ ID NOS: 87 and 88, SEQ ID NOS: 89 and 90, SEQ ID NOS: 91 and 92, SEQ ID NOS: 93 and 94, SEQ ID NOS: 95 and 96, SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100, SEQ ID NOS: 101 and 102, SEQ ID NOS: 103 and
104. DATED this 17 h day of April, 2001 SE. I. DU PONT DE NEMOURS AND COMPANY By their Patent Attorneys: CALLINAN LAWRIE
AU65751/98A 1997-03-24 1998-03-19 A method to identify and breed corn with increased kernel oil concentration Ceased AU734755B2 (en)

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US20040025202A1 (en) * 2002-03-15 2004-02-05 Laurie Cathy C. Nucleic acid molecules associated with oil in plants
WO2007103738A2 (en) * 2006-03-01 2007-09-13 Pioneer Hi-Bred International, Inc. Compositions related to the quantitative trait locus 6 (qtl6) in maize and methods of use
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