AU6575198A - 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

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AU6575198A
AU6575198A AU65751/98A AU6575198A AU6575198A AU 6575198 A AU6575198 A AU 6575198A AU 65751/98 A AU65751/98 A AU 65751/98A AU 6575198 A AU6575198 A AU 6575198A AU 6575198 A AU6575198 A AU 6575198A
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Robert Stefan Reiter
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EIDP Inc
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EI Du Pont de Nemours and Co
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/13—Plant traits
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

Description

TITLE
A METHOD TO IDENTIFY AND BREED CORN WITH
INCREASED KERNEL OIL CONCENTRATION
FIELD OF INVENTION The invention 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; i.e., 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 time- consuming and an expensive labor. 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 (i.e., 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. (5_ :103-111) and livestock (Nordstrom, J.W. et al. (1972) J. An. Sci 3.5 (2,1:357-361). 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 57:1-7) 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. 40th Ann. Corn and Sorghum Res. Conf. Am. Seed Trade Assoc, Washington D.C. pp. 66-89) measured isozyme allelic frequency changes following twenty- five 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, I.L., 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 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 associated with increased oil. Some of the regions identified by significant R LP marker loci may be in common between the two studies, however of the fifteen RJLP 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 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 one or more genetic markers to select a corn plant from a corn breeding population by marker- assisted selection, wherein the genetic markers are selected from the group consisting of sl375, sl384, sl394, sl416, sl422, sl432, sl457, sl480, sl476, sl478, sl484, sl500, sl513, sl529, sl544, sl545, sl630, sl633, sl647, sl750, sl756, sl757, sl767, sl772, sl774, sl780, sl797, sl813, sl816, sl817, sl836, sl853, sl860, sl870, sl921, sl922, sl925, sl931, sl933, sl939, sl946, sl949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175; 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 high oil corn germplasm is a member of an Alexho synthetic population or a progeny thereof.
Also disclosed is a method for identifying com plants or com lines for use as parents for creation of a breeding population, the method comprising a) genotyping com plants or com lines with one or more genetic markers wherein the genetic markers are selected from the group consisting of sl375, sl384, sl394, sl416, S1422, sl432, sl457, sl480, sl476, sl478, sl484, sl500, sl513, sl529, sl544, sl545, sl630, sl633, sl647, sl750, sl756, sl757, sl767, sl772, sl774, sl780, sl797, sl813, sl816, sl817, sl836, sl853, sl860, sl870, sl921, sl922, sl925, sl931, sl933, sl939, sl946, sl949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175; and b) identifying com plants or com 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 com kernel oil concentration. These oil alleles were initially identified in materials composed of or derived from the Alexho synthetic breeding populations. Further, the method facilitates the use of this high oil material in breeding programs with the objective of developing new high oil com germplasm.
Specifically, the method uses genetic markers to predict the oil breeding value of lines in a com 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 association with kernel oil concentration. The method is only limited by the ability to measure polymorphism at a given marker locus. Those skilled in the art will recognize 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 si 375, si 384, sl394, sl416, sl422, sl432, sl457, sl476, sl478, sl480, sl484, sl500, sl513, S1529, sl544, sl545, sl630, sl633, sl647, sl750, sl756, sl757, sl767, sl772, sl774, sl780, sl797, sl813, sl816, sl817, sl836, sl853, sl860, sl870, sl921, sl922, S1925, sl931, sl933, sl939, sl946, sl949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175. A further embodiment of the present invention are the trait loci controlling the expression of com kernel oil concentration. These loci are identified and defined (i.e., mapped) by the marker loci of the present invention.
An additional embodiment of the present invention are com plants and high oil com 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 com 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 com kernel oil concentration
Marker Sequence (5'-3') Primer Type SEQ ID NO. sl375 TTTATGGGTTGGGAGATACTTG forward 1
AGATGTGTGCGTTTTTGAGAG reverse 2 sl384 TTACGGCCTAGACATTTCGAC forward 3
CACTTGCTTTCAGGTACCCA reverse 4 sl394 CTGCCCAGTCCGTAATGAA forward 5
TAGATTTATTTTCTGAACGATTGG reverse 6 sl416 GATCTCTCTGAGGCTTGTCC forward 7
TGTAGTTGAGGATGCTCCC reverse 8 si 422 AGGCAAGGCTTTCTTCATAC forward 9
CGGACGACGACTGTGTTC reverse 10 si 432 ACATGAGAAACAAGATAGAACCAG forward 11
AAAATGTAAGAACTTGTTTGGGA reverse 12 sl457 CTGCTTATTGCTTTCGTCATA forward 13 TGCTGCACTACTTGAACCTAG reverse 14 si 476 ACACAGAGATGACAAAAGCAA forward 15 GCAGGCGTGCTATGAGAG reverse 16 si 478 AGCGGTGAAACCCTTATG forward 17 CTGTGGCTGGTTCCTCTC reverse 18 si 480 GCTCTTGATAAAAAGGCAAGT forward 19 CTTGTTGTAATGGATGAGTGAG reverse 20 si 484 GCTCGTAGTAGGGGTTACG forward 21 GACAGCCTCACCTCAAGA reverse 22 s l500 ACAGATCTTGACACGTACATACC forward 23 GGACGTGTATCCTCAAATCAT reverse 24 sl 513 CAGCGAATACTGAATAACGC forward 25 TGTTGGATGAGCACTGAAC reverse 26 si 529 TGTTCTCAACAACCACCG forward 27
_ _ CGTTTAGCGATATCATTTTCC reverse 28 si 544 GATCCTACCAAAATCTTATAGGC forward 29
ACAGCTAGCCAAGATCTGATT reverse 30 sl545 CGATACTAATGGAAGCCCTAA forward 31 ATGGCCCATTAAGTTTATCAC reverse 32
S1630 AAAGCGTAGTCGGAAAGC forward 33 ACCAATGATCTTTACGCAGAT reverse 34 sl633 TAATCAGAGCGTACATCAGGA forward 35
AGGGCATCAATCAAGAATG reverse 36 si 647 GAGACTTTTGAGGAGAAAGCA forward 37
GATCAAAAGAGCAAAAGGAGA reverse 38 si 750 AACTGATGAATACCTTCCCAG forward 39
TGATTAACTTCTCCCTTTGGT reverse 40 sl756 TCGGCACAACATATGAGTTAC forward 41
CCCCCATAGAGAGAGATAGAG reverse 42 sl757 AAGCACGGCCCAATAGAAT forward 43
AGGATGTCCCTAGCTTTATTG reverse 44 si 767 TCATTGCCCAAAGTGTTG forward 45
CTCATCACCCCTCCAGAG reverse 46 si 772 GATCCACGCCATTTAAAC forward 47
TGATACTCTGGTGCATGTTC reverse 48 si 774 GATCGCTCCGATCTATCC forward 49
AGCGGCATCTATGTTCTATG reverse 50 si 780 CCCAGTGCGAAGAGACTC forward 51 ACACCTGCTCTGCACCAC reverse 52 si 797 CTAACCCACGACGACCCT forward 53 GCATGAGTGCATGTGCAT reverse 54 sl813 CTGCCACATGCTTTTCTG forward 55 CTGTAAAGAAGCTGGTCTGGA reverse 56 sl816 TTCTCCTCATGGATGCGT forward 57 CTATTTGGAAGTATGGGCTTCA reverse 58 sl817 GAGGGCATCTATGTGCAAC forward 59
GCTCAGAAGTTGCGTTTATG reverse 60 sl836 TTCCTTCACGTTTCTCTGTTAA forward 61
CACATAAACCTAATGGGGTACA reverse 62 sl853 CCCAAAGGCGATACCTATT forward 63
CCCACTTTCTCACTCTTTTCT reverse 64 sl860 GAGGTGAGTACTATGCAAATGC forward 65
CAGGCTTACCTAGCCTTCTC reverse 66 sl870 CTATGGATGGCTGCTTGC forward 67
GTCAGGCAGCAGAATGTG reverse 68 sl921 AAACCGTCCAGCGACTAC forward 69
GGAAGAACCAATCCCATATCT reverse 70 si 922 AACATCCTGTCGGAAACAG forward 71
TCATCACGTCTCTCTTTCAAC reverse 72 si 925 TTGTGGCAGAATCTCAAATTA forward 73
CGACTGGTGACATGTGAAG reverse 74 sl931 AGTGAGGAAAGAATATGCTGG forward 75
TGGACTGAGAAACTGATTTGA reverse 76 sl933 CACAAATGTGAAGGTAAACACT forward 77
AATGGTACGGTTCAGGATG reverse 78 sl939 AGATGACGCACGGAACAC forward 79
AGCATCATGTAGCAGGAGG reverse 80 si 946 TTGCAGCACTGTCGTAGTC forward 81
GCGCGAGTGGAGTAGTAAG reverse 82 si 949 AAGATTATGCAGATGAGACACC forward 83
GTTCCATGCTTTCCTTGG reverse 84 s2054 GCCGATACCATGTAAGAGAAT forward 85
CTCTGGGCTCTGTGTTAGAGT reverse 86 s2055 CTGCTTTCTCTGTTCCAGC forward 87 AATCGCTTACTTGTAACCCAC reverse 88 s2057 AAGAACGTACGTCCCATAAAG forward 89 CAAGGTAAAGTGACAAAGCAG reverse 90 s2058 GTTCAGGATGAGGCGGAA forward 91 GTGATCATCGCAGGAGACC reverse 92 s2097 GGAGCCTGGAGTGAGAAC forward 93 CATGCTCACCTAACGTGG reverse 94 s2122 ATCTGAACACTTGAGCAACAA forward 95 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: Com. Any variety, cultivar, or population of Zea mavs 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 Com Germplasm. This term characterizes com 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 com germplasm include but are not limited to open-pollinated varieties, hybrids, synthetics, inbred lines, races, and populations or com 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. 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 com germplasm developed by Denton Alexander at the University of Illinois. Alexho synthetic high oil com 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 polymoφhism. Examples of genetic markers includes but is not limited to RPLPs, RAPDs, allozymes, SSRs, and AFLPs.
Marker locus. The genetically defined location of DNA polymoφhisms 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 Polymoφhism (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 Polymoφhic DNA (RAPD). A DNA amplification- based genetic marker in which short, sequence arbitrary primers are used and the resulting amplification products are size separated and differences in amplification patterns observed (Williams J.G.K. et al. 1990. Nucleic Acids Res. J 8:6531-6535).
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. 7/2:4127-4138).
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 com 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 self- pollinated 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 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 com 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 kemels are highly represented in ASKC28 and as such I have designated the ASKC28 as being waxy). The FI plants were selfed and resulting F2 populations were grown. Individual F2 plants were selfed and derived kemels were advanced using single seed decent through six generations of selfing (S6) to produce recombinant inbred lines. Up to twenty kemels 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 (LH119wx 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. 5:4321-4325). DNA samples were quantified via fluorescence using YoPro-1™ iodide (Molecular Probes, Inc., Eugene, OR) and diluted to 4 μg/ml.
SSR regions for each DNA sample were analyzed using the following protocol: 1. Ten μl of amplification cocktail (see Table 2) was added to 5 μl
(20 ng) of extracted DNA;
2. The DNA fragment flanked by sequences complementary to the primers present in the amplification cocktail was amplified by PCR (U.S. Patent No. 4,683,202 and U.S. Patent No. 4,683,195) using the following protocol: 1) 45 cycles of 50 sec at 95°C, 50 sec at 54°C and 80 sec at 72°C and 2) 1 cycle of 300 sec at 72°C;
3. Approximately 8 μl of each sample was loaded onto agarose gels composed of 2% Metaphor (FMC Corp., Rockland, ME), IX TBE, and 0.5 μg/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 μg/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 1.5X dNTPs 2 mM 0.3 mM
Forward Primer 40 μM 0.45 μM
Reverse Primer 40 μM 0.45 μM
AmpliTaq Polymerase™ 5 U/μl 0.05 U/μl
* 10X Buffer is a pH 9.0 solution composed of 800 mM Tris-OH, 200 mM
(NH4)2SO4, and 25 mM MgCl2.
Localization of oil loci
One hundred thirty three polymoφhic SSR marker loci were used to genotype the recombinant inbreds from the LH119wx x ASKC28wx cross and one hundred and three polymoφhic SSR marker loci were used to genotype the LH51 x ASKC28wx-derived population. In addition, twenty publicly available polymoφhic 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 S.E., et al. (1993) Whitehead Inst. Biomed. Res., Cambridge, MA). This resulted in the establishment often 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 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, M.D., 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 LH119wx 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 sl922
B 1 sl478, sl853,sl949
C 1 S1860, sl925, sl931,s2150
D 2 s2175 E si 394
F 4 sl476, sl772, sl816, s2122, sl836
G 4 sl939, sl946
H 4 sl870
I 5 sl529
J 5 s2054, sl647, sl500, sl545, sl774, s2097
K 6 sl457, s2055, sl757, s2125, sl780, sl375, sl797, sl416, sl432, sl921
L 7 sl630, sl422, s2156
M 8 sl817, s2057
N 9 sl544, sl633, sl384, sl813, sl767, s2058, sl933, sl513, sl484
O 10 sl756
P 10 si 480 (positive oil allele in LH51)
Q N.A.* sl750
*N.A. - chromosome location not known
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 LH119wx x ASKC28wx had a positive oil effect versus the alternative LH119- derived 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 si 480, 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 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 com 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 com inbreds. The resultant hybrid could be self-pollinated to produce an F2 population for the puφoses 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. 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 com 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 (i.e., 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 (A-E), 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.
SEQUENCE LISTING GENERAL INFORMATION :
( l ) APPLICANT :
(A) ADDRESSEE: E. I. DU PONT DE NEMOURS AND COMPANY
(B) STREET: 1007 MARKET STREET
(C) CITY: WILMINGTON
(D) STATE: DELAWARE
(E) COUNTRY: USA
(F) ZIP: 19898
(G) TELEPHONE: 302-992-4926 (H) TELEFAX: 302-773-0164 (I) TELEX: 6717325
(li) TITLE OF INVENTION: A METHOD TO IDENTIFY AND BREED
CORN WITH INCREASED KERNEL OIL CONCENTRATION
(ill) NUMBER OF SEQUENCES: 104
(iv) COMPUTER READABLE FORM:
(A) MEDIUM TYPE DISKETTE, 3.50 INCH
(B) COMPUTER: IBM PC COMPATIBLE
(C) OPERATING SYSTEM: MICROSOFT WINDOWS 95
(D) SOFTWARE: MICROSOFT WORD VERSION 7.0A
(v) CURRENT APPLICATION DATA:
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(B) FILING DATE:
(C) CLASSIFICATION:
(vi) PRIOR APPLICATION DATA:
(A) APPLICATION NUMBER: 60/041,515
(B) FILING DATE: MARCH 24, 1997
(C) CLASSIFICATION:
(vii) ATTORNEY/AGENT INFORMATION:
(A) NAME: MAJARIAN, WILLIAM R.
(B) REGISTRATION NUMBER: P-41,173
(C) REFERENCE/DOCKET NUMBER: BB-1076
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(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:51: CCCAGTGCGA AGAGACTC 18
(2) INFORMATION FOR SEQ ID NO:52:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 52: ACACCTGCTC TGCACCAC 18 (2) INFORMATION FOR SEQ ID NO: 53:
(l) SEQUENCE CHARACTERISTICS.
(A) LENGTH: 18 base pairs
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(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
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(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 53: CTAACCCACG ACGACCCT 18
(2) INFORMATION FOR SEQ ID NO: 54:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
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(xi) SEQUENCE DESCRIPTION: SEQ ID NO:54: GCATGAGTGC ATGTGCAT 18
(2) INFORMATION FOR SEQ ID NO: 55:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
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(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 55: CTGCCACATG CTTTTCTG 18
(2) INFORMATION FOR SEQ ID NO:56:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 56: CTGTAAAGAA GCTGGTCTGG A 21 (2) INFORMATION FOR SEQ ID NO: 57:
( ) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 57: TTCTCCTCAT GGATGCGT 18
(2) INFORMATION FOR SEQ ID NO: 58:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 58: CTATTTGGAA GTATGGGCTT CA 22
(2) INFORMATION FOR SEQ ID NO: 59:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
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(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 59: GAGGGCATCT ATGTGCAAC 19
(2) INFORMATION FOR SEQ ID NO: 60:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 20 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid (x ) SEQUENCE DESCRIPTION: SEQ ID NO: 60: GCTCAGAAGT TGCGTTTATG 20 (2) INFORMATION FOR SEQ ID N0:61.
(1) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 61: TTCCTTCACG TTTCTCTGTT AA 22
(2) INFORMATION FOR SEQ ID NO: 62:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 62: CACATAAACC TAATGGGGTA CA 22
(2) INFORMATION FOR SEQ ID NO: 63:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
( i) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 63: CCCAAAGGCG ATACCTATT 19
(2) INFORMATION FOR SEQ ID NO: 64:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 64: CCCACTTTCT CACTCTTTTC T 21 (2) INFORMATION FOR ΞEQ ID NO: 65:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
( ) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 65: GAGGTGAGTA CTATGCAAAT GC 22
(2) INFORMATION FOR SEQ ID NO: 66:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 20 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 66: CAGGCTTACC TAGCCTTCTC 20
(2) INFORMATION FOR SEQ ID NO: 67:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 67: CTATGGATGG CTGCTTGC 18
(2) INFORMATION FOR SEQ ID NO: 68:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 68: GTCAGGCAGC AGAATGTG 18 (2) INFORMATION FOR SEQ ID NO: 69:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic ac d
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 69: AAACCGTCCA GCGACTAC 18
(2) INFORMATION FOR SEQ ID NO: 70:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 70: GGAAGAACCA ATCCCATATC T 21
(2) INFORMATION FOR SEQ ID NO: 71:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(Xi) SEQUENCE DESCRIPTION: SEQ ID NO: 71: AACATCCTGT CGGAAACAG 19
(2) INFORMATION FOR SEQ ID NO: 72:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 72: TCATCACGTC TCTCTTTCAA C 21 (2) INFORMATION FOR SEQ ID NO:73:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
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(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
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(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 73: TTGTGGCAGA ATCTCAAATT A 21
(2) INFORMATION FOR SEQ ID NO: 74:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 74: CGACTGGTGA CATGTGAAG 19
(2) INFORMATION FOR SEQ ID NO: 75:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 75: AGTGAGGAAA GAATATGCTG G 21
(2) INFORMATION FOR SEQ ID NO: 76:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 76: TGGACTGAGA AACTGATTTG A 21 (2) INFORMATION FOR SEQ ID NO: 77:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 77: CACAAATGTG AAGGTAAACA CT 22
(2) INFORMATION FOR SEQ ID NO: 78:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 78: AATGGTACGG TTCAGGATG 19
(2) INFORMATION FOR SEQ ID NO: 79:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:79: AGATGACGCA CGGAACAC 18
(2) INFORMATION FOR SEQ ID NO:80:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:80: AGCATCATGT AGCAGGAGG 19 (2) INFORMATION FOR SEQ ID NO: 81
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
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(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 81: TTGCAGCACT GTCGTAGTC 19
(2) INFORMATION FOR SEQ ID NO: 82:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:82: GCGCGAGTGG AGTAGTAAG 19
(2) INFORMATION FOR SEQ ID NO:83:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
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(n) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 83: AAGATTATGC AGATGAGACA CC 22
(2) INFORMATION FOR SEQ ID NO: 84:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
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(C) STRANDEDNESS: single
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(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 84: GTTCCATGCT TTCCTTGG 18 (2) INFORMATION FOR SEQ ID NO:85:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
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(xi) SEQUENCE DESCRIPTION: SEQ ID NO:85: CTCTGGGCTC TGTGTTAGAG T 21
(2) INFORMATION FOR SEQ ID NO: 86:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:86: CTCTGGGCTC TGTGTTAGAG T 21
(2) INFORMATION FOR SEQ ID NO: 87:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 87: CTGCTTTCTC TGTTCCAGC 19
(2) INFORMATION FOR SEQ ID NO: 88:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO:88: AATCGCTTAC TTGTAACCCA C 21 (2) INFORMATION FOR SEQ ID NO:89:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 89: AAGAACGTAC GTCCCATAAA G 21
(2) INFORMATION FOR SEQ ID NO: 90:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 90: CAAGGTAAAG TGACAAAGCA G 21
(2) INFORMATION FOR SEQ ID NO: 91:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 91: GTTCAGGATG AGGCGGAA 18
(2) INFORMATION FOR SEQ ID NO: 92:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 19 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 92: GTGATCATCG CAGGAGACC 19 (2) INFORMATION FOR SEQ ID NO: 93
(i) SEQUENCE CHARACTERISTICS.
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
( i) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 93: GGAGCCTGGA GTGAGAAC 18
(2) INFORMATION FOR SEQ ID NO: 94:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 94: CATGCTCACC TAACGTGG 18
(2) INFORMATION FOR SEQ ID NO: 95:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 95: ATCTGAACAC TTGAGCAACA A 21
(2) INFORMATION FOR SEQ ID NO: 96:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 96: ATAGACCGGA CCCATCAC 18 (2) INFORMATION FOR SEQ ID NO: 97:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(n) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 97: CGAACAGCGG GTACACCT 18
(2) INFORMATION FOR SEQ ID NO: 98:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 20 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 98: GAGGTCAGCT TCCTCGATCT 20
(2) INFORMATION FOR SEQ ID NO: 99:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 99: GGAATCGTTC CTCCACAC 18
(2) INFORMATION FOR SEQ ID NO: 100:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 100: CTTCCTCGGT GTCAGACG 18 (2) INFORMATION FOR SEQ ID NO: 101:
( ) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 101: ATGGAAACAT CAAAGTGGAT T 21
(2) INFORMATION FOR SEQ ID NO: 102:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 20 base pairs
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(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 102: TGCTACCCTG ATGACCTGAT 20
(2) INFORMATION FOR SEQ ID NO: 103:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid
(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 103: ACCACTAGTC TCATATGAAG GG 22
(2) INFORMATION FOR SEQ ID NO: 104:
(l) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 18 base pairs
(B) TYPE: nucleic acid
(C) STRANDEDNESS: single
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: other nucleic acid (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 104: GGTAGGTGGG TAGGGGTT 18

Claims (20)

What is claimed:
1. A method of breeding for com with increased kemel oil concentration comprising: a) using one or more genetic markers to select a com plant from a com breeding population by marker-assisted selection, wherein the genetic markers are selected from the group consisting of sl375, sl384, sl394, sl416, sl422, sl432, sl457, sl480, sl476, sl478, sl484, sl500, sl513, sl529, sl544, sl545, sl630, sl633, sl647, sl750, sl756, sl757, sl767, sl772, sl774, sl780, sl797, sl813, sl816, sl817, sl836, sl853, sl860, sl870, sl921, sl922, sl925, sl931, sl933, sl939, sl946, sl949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175; and b) crossing the selected com plant with a second com plant wherein the progeny com plants of the cross display increased kemel oil concentration.
2. The method of claim 1 wherein the selected com plant is member of an Alexho synthetic population or a progeny thereof.
3. A method for identifying com plants or com lines for use as parents for creation of a breeding population, the method comprising: a) genotyping com plants or com lines with one or more genetic markers wherein the genetic markers are selected from the group consisting of sl375, sl384, sl394, sl416, sl422, sl432, sl457, sl480, sl476, sl478, sl484, sl500, sl513, sl529, sl544, sl545, sl630, sl633, sl647, sl750, sl756, sl757, sl767, sl772, sl774, sl780, sl797, sl813, sl816, sl817, sl836, sl853, sl860, sl870, sl921, sl922, sl925, sl931, sl933, sl939, sl946, sl949, s2054, s2055, s2057, s2058, s2097, s2122, s2125, s2150, s2156 and s2175; and b) identifying com plants or com lines which, based upon their genotype, are predicted to produce transgressive segregants for kemel oil concentration.
4. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s2054, si 647, si 500, si 545, sl774 and s2097.
5. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of si 817 and s2057.
6. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of si 860, si 931, s2150 and sl925.
7. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of sl457, s2055, sl757, s2125, sl780, sl375, sl797, si 416, sl432 and sl921.
8. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of sl544, sl633, sl384, sl813, sl767, s2058, sl933, sl513 and sl484.
9. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of si 476, si 772, si 816, s2122 and sl836.
10. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of si 939 and si 946.
11. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of sl478, sl853 and sl949.
12. A trait locus controlling kemel oil concentration, the locus mapped by a genetic marker selected from the group consisting of s 1630, s 1422 and s2156.
13. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker si 756.
14. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker si 922.
15. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker si 529.
16. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker sl394.
17. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker s 1750.
18. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker si 870.
19. A trait locus controlling kemel oil concentration, the locus mapped by the genetic marker s2175.
20. Com plants that display increased kemel oil concentration produced by the method of Claim 1.
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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Families Citing this family (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6806399B1 (en) 2000-04-19 2004-10-19 Carmel-Haifa University Economic Corporation Ltd. Pollen-mediated method for transformation of maize, tomato or melon
BRPI0206998B1 (en) * 2001-02-06 2015-08-18 Dow Agrosciences Llc High oil level genetic marker assisted selection method in maize plants.
US20040025202A1 (en) * 2002-03-15 2004-02-05 Laurie Cathy C. Nucleic acid molecules associated with oil in plants
CA2638739A1 (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
WO2007101273A2 (en) * 2006-03-01 2007-09-07 Pioneer Hi-Bred International, Inc. Compositions related to the quantitative trait locus 6 (qtl6) in maize and methods of use
WO2009089928A1 (en) * 2008-01-18 2009-07-23 Syngenta Participations Ag Maize plants characterised by quantitative trait loci (qtl)
EP1947198A1 (en) * 2007-01-18 2008-07-23 Syngeta Participations AG Maize plants characterised by quantitative trait loci (QTL)
JP2008220269A (en) * 2007-03-13 2008-09-25 Japan Grassland Farming Forage Seed Association Primer set for detecting dna marker linked to gene locus relating to fat-content in corn seed and its use

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5492547B1 (en) * 1993-09-14 1998-06-30 Dekalb Genetics Corp Process for predicting the phenotypic trait of yield in maize
US5476524A (en) * 1993-12-09 1995-12-19 Zeneca Limited Oil producing sunflowers and production thereof

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AU734755B2 (en) 2001-06-21
WO1998042870A1 (en) 1998-10-01
JP2001517951A (en) 2001-10-09
NZ337906A (en) 2001-08-31
HUP0001745A3 (en) 2002-04-29
KR20010005625A (en) 2001-01-15
AR012152A1 (en) 2000-09-27
CA2280933A1 (en) 1998-10-01
IL131908A0 (en) 2001-03-19
HUP0001745A2 (en) 2000-09-28
PL335910A1 (en) 2000-05-22
EP0972079A1 (en) 2000-01-19

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