New corn plants

FIELD: agriculture.

SUBSTANCE: during selection by markers, a set of alleles related to loci of quantity tokens (QTL) that contribute to expression of certain phenes of economic value is introduced into a corn idioplasm. The criteria are selected among grain crop capacity, grain moisture when picked, early and late root fit, stem drowning, frequency of common rust, frequency of ear rotting caused by Fusarium (ear wilt), resistance to Sulcotrione and panicle structure. Invention also relates to the method of such plants production, and also to methods of analysis and screening to identify plants with a required profile of alleles.

EFFECT: improved method of new corn plants production.

33 cl, 1 dwg, 19 tbl

 

The text descriptions are given in facsimile form.

1. Plant corn containing the nuclear genome, including a set of favourable alleles at the corresponding set of at least 14 loci QTL, each of which contributes to the phenotypic trait of grain yield, and:
(a) each locus QTL gene is automatically concatenated with at least one marker locus, selected from the group of loci characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 59/60 and 77/78, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 77/78 and 27/28, respectively, identifying a pair of markers linked to QTL2;
SEQ ID NO: 47/48 and 75/76, respectively, identifying a pair of markers linked to QTL3;
SEQ ID NO: 65/66 and 9/10, respectively, identifying a pair of markers linked to QTL4;
SEQ ID NO: 69/70 and 13/14, respectively, identifying a pair of markers linked to QTL5;
SEQ ID NO: 73/74 and 25/26, respectively, identifying a pair of markers linked to QTL6;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL7;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL8;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL9;
SEQ ID NO: 41/42 and 49/50, respectively, identifying a pair of markers linked to QTL10;
SEQ ID NO: 49/50 and 61/62, respectively, identifying a pair of markers linked to QTL11;
SEQ ID NO: 17/18 and 51/52, respectively, identifying a pair of markers linked to QTL12;
SEQ ID NO: 51/5 and 19/20, accordingly, identifying a pair of markers linked to QTL13; and
SEQ ID NO: 29 and 30, identifying a marker linked with QTL14; and
b) each allele at the corresponding QTL locus is determined by the product of PCR amplification, which is essentially identical to the corresponding amplification product of the favourable allele specified in table, which can be obtained from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction using a pair of primers identified in paragraph (a).

2. Plant corn according to claim 1, which contains a full set of favourable alleles at the corresponding 14 loci QTL.

3. Plant corn according to claim 1, in which:
the QTL loci 1-4 localized on chromosome 1,
the QTL loci 5 and 6 are localized on chromosome 2,
the loci QTL 7-9 localized on chromosome 4,
the loci QTL 10-13 localized on chromosome 5,
the QTL locus 14 is localized on chromosome 7.

4. Plant corn in any one of claims 1 to 3, further comprising 11 QTL that contribute in the moisture content of the grain, which are mapped to loci on chromosomes 1, 2, 3, 4, 5, 7 and 8, with each allele at the corresponding QTL is defined by at least one marker allele at the specified at least one marker locus linked to the QTL and the marker allele is characterized by the product of PCR amplification with the respective pair of oligonucleotide primers, lead the military in the table, moreover, the amplification product is essentially identical to the corresponding amplification product of the favourable allele indicated in tables A-G, which can be obtained from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction with identical pairs of primers.

5. Plant corn according to claim 4, in which for QTL contributing in the moisture content of the grain:
(a) each locus QTL is genetically linked to at least one marker locus, which is characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 23/24 and 3/4, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 65/66 and 9/10, respectively, identifying a pair of markers linked to QTL2;
SEQ ID NO: 69/70 and 13/14, respectively, identifying a pair of markers linked to QTL3;
SEQ ID NO: 71/72 and 53/54, respectively, identifying a pair of markers linked to QTL4;
SEQ ID NO: 53/54 and 57/58, respectively, identifying a pair of markers linked to QTL5;
SEQ ID NO: 43/44, identifying a marker linked to QTL6;
SEQ ID NO: 5/6 and 37/38, respectively, identifying a pair of markers linked to QTL7;
SEQ ID NO: 21/22 and 33/34, respectively, identify them a couple of markers, coupled with QTL8;
SEQ ID NO: 31/32 and 39/40, respectively, identifying a pair of markers linked to QTL9;
SEQ ID NO: 29/30, identifying a marker linked with QTL10;
SEQ ID NO: 67/68, identifying a marker linked with QTL11; and
b) each allele at the corresponding QTL locus is determined by the product of PCR amplification, which is essentially identical to the corresponding amplification product of the favourable allele specified in tablw, which can be obtained from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction using a pair of primers identified in paragraph (a).

6. Plant corn according to claim 5, in which:
the QTL loci 1 and 2 are localized on chromosome 1,
the loci QTL 3-5 localized on chromosome 2,
locus QTL 6 is localized on chromosome 3,
locus QTL 7 is localized on chromosome 4,
locus QTL 8 is localized on chromosome 5,
the loci QTL 9 and 10 are localized on chromosome 7, and
locus QTL 11 is localized on chromosome 8.

7. Plant corn according to claim 6, which contains a full set of favourable alleles at the corresponding set of 11 loci qtls contributing to the phenotypic trait of grain moisture in the collection.

8. Plant corn in any one of claims 1 to 3, containing at least one additional set of favourable alleles at the corresponding loci QTL contributing to the basal and stableboy lodging, and these QTL genetic the Eski coupled with at least one additional marker locus, selected from the group of marker loci, characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 3/4, and 59/60, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 27/28 and 47/48, respectively, identifying a pair of markers linked to QTL2;
SEQ ID NO: 45/46, identifying a marker linked with QTL3.

9. Plant corn in claim 8, in which the QTL loci 1, 2 and 3 are localized on chromosome 1.

10. Plant corn in any one of claims 1 to 3, containing at least one additional favorable allele at the corresponding QTL locus contributing to the frequency smut ordinary, and QTL is genetically linked to at least one additional marker locus, characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in SEQ ID NO: 11/12, identifying a marker linked to QTL1.

11. Plant corn in claim 10, in which the locus QTL1 is localized on chromosome 3.

12. R is stenie corn according to any one of claims 1 to 3, containing at least one additional set of favourable alleles at the corresponding loci QTL contributing to the structure of the panicle, and these QTL is genetically linked to at least one additional marker locus selected from the group of marker loci, characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 11/12, identifying a marker linked to QTL1;
SEQ ID NO: 55/56, identifying a marker linked with QTL2;
SEQ ID NO: 31/32 and 39/40, respectively, identifying a pair of markers linked to QTL3;
SEQ ID NO: 81/82 and 7/8, respectively, identifying a pair of markers linked to QTL4.

13. Plant corn at para.12, which:
locus QTL1 is localized on chromosome 3,
locus QTL2 is localized on chromosome 6,
locus QTL3 is localized on chromosome 7, and
locus QTL4 is localized on chromosome 9.

14. Plant corn in any one of claims 1 to 3, containing at least one additional set of favourable alleles at the corresponding loci QTL contributing to the resistance to Sulcotrione, and these QTL is genetically linked to at least one additional marker locus that is selected from the group of marker loci, characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 43/44, identifying a marker linked to QTL1;
SEQ ID NO: 81/82 and 7/8, respectively, identifying a pair of markers linked to QTL2.

15. Plant corn at 14, which:
locus QTL1 is localized on chromosome 3, and
locus QTL2 is localized on chromosome 9.

16. Plant corn in any one of claims 1 to 3, containing at least one additional set of favourable alleles at the corresponding loci QTL contributing to the resistance to Fusarium head blight, and these QTL is genetically linked to at least one additional marker locus selected from the group of marker loci, characterized by at least one pair of linked markers each of which can be identified by using a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer having the nucleotide sequence shown in:
SEQ ID NO: 1/2 and 79/80, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 79/80 and 15/16, respectively, identifying a pair of markers linked to QTL2.

17. Plant corn in clause 16, in which the QTL loci 1 and 2 are localized on chromosome 5.

18. Plant corn in any one of claims 1 to 3, which always contains the most favorable allele at a marker locus linked to the QTL, and/or show the indicator LOT, given in table-G.

19. Plant corn on p, which contains at least one copy of the most favorable alleles at each locus.

20. Plant corn according to claim 1, which is an inbred plant.

21. Plant corn according to claim 1, which is a hybrid, especially F1-a hybrid from a single crossing.

22. Plant corn according to claim 1, in which at least part of these loci QTL derived from inbred lines M/2 and M/1, respectively, deposited at NCIMB under the number NCIMB 41460 and NCIMB 41459.

23. Plant material, including plant parts, plant seeds, and products of corn processing, in particular grain and the seeds of corn derived from a plant according to any one of claims 1 to 22.

24. Products of corn processing, in particular grain and the seeds of corn derived from a plant according to any one of claims 1 to 22.

25. The set of markers comprising a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer capable of identifying a marker linked to the QTL locus contributing to fruitful is to be grain, these primers have the nucleotide sequence shown in:
SEQ ID NO: 59/60 and 77/78, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 77/78 and 27/28, respectively, identifying a pair of markers linked to QTL2;
SEQ ID NO: 47/48 and 75/76, respectively, identifying a pair of markers linked to QTL3;
SEQ ID NO: 65/66 and 9/10, respectively, identifying a pair of markers linked to QTL4;
SEQ ID NO: 69/70 and 13/14, respectively, identifying a pair of markers linked to QTL5;
SEQ ID NO: 73/74 and 25/26, respectively, identifying a pair of markers linked to QTL6;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL7;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL8;
SEQ ID NO: 35/36 and 63/64, respectively, identifying a pair of markers linked to QTL9;
SEQ ID NO: 41/42 and 49/50, respectively, identifying a pair of markers linked to QTL10;
SEQ ID NO: 49/50 and 61/62, respectively, identifying a pair of markers linked to QTL11;
SEQ ID NO: 17/18 and 51/52, respectively, identifying a pair of markers linked to QTL12;
SEQ ID NO: 51/52 and 19/20, respectively, identifying a pair of markers linked to QTL13;
SEQ ID NO: 29 and 30, identifying a marker linked with QTL14;
moreover, these primers in the PCR reactions give an amplification product exhibiting such molecularlevel or nucleotide sequence, which is essentially identical to the corresponding product of PCR amplification, which can be obtained from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction with identical pairs of primers.

26. The set of markers on A.25, additionally including a pair of oligonucleotide primers for PCR, consisting of direct primer and a reverse primer capable of identifying a marker linked to the QTL locus contributing in the moisture content of the grain in the collection, and these primers have the nucleotide sequence shown in:
SEQ ID NO: 23/24 and 3/4, respectively, identifying a pair of markers linked to QTL1;
SEQ ID NO: 65/66 and 9/10, respectively, identifying a pair of markers linked to QTL2;
SEQ ID NO: 69/70 and 13/14, respectively, identifying a pair of markers linked to QTL3;
SEQ ID NO: 71/72 and 53/54, respectively, identifying a pair of markers linked to QTL4;
SEQ ID NO: 53/54 and 57/58, respectively, identifying a pair of markers linked to QTL5;
SEQ ID NO: 43/44, identifying a marker linked to QTL6;
SEQ ID NO: 5/6 and 37/38, respectively, identifying a pair of markers linked to QTL7;
SEQ ID NO: 21/22 and 33/34, respectively, identifying a pair of markers linked to QTL8;
SEQ ID NO: 31/32 and 39/40, respectively, identifying a pair of markers linked to QTL9;
SEQ ID NO: 29/30, identifying a marker linked with QL10;
SEQ ID NO: 67/68, identifying a marker linked with QTL11;
moreover, these primers in the PCR reactions give an amplification product having such a molecular weight or a nucleotide sequence that is essentially identical to the corresponding product of PCR amplification, which can be obtained from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction with identical pairs of primers.

27. The manner of identification of maize that contains many of the most favorable alleles at marker loci linked with the corresponding loci QTL, which includes the following stages:
i) obtaining plant material from the studied plant or population of plants and extraction of DNA from this material;
ii) analysis of the DNA sample obtained in stage i)to determine the allelic variants available for a marker loci genetically linked with the corresponding qtls contributing to the phenotypic trait selected from the group of grain yield and grain moisture at harvest, by:
(a) applying a set of markers for p. 25 or 26 when reaction PCR amplification;
b) identification of marker alleles by determining the molecular weights and/or the nucleotide sequences of PCR products amplification obtained in stage a);
c) comparing the identified at stage b) forefront of the popular scales and/or the nucleotide sequences of PCR products amplification with molecular weights and/or the nucleotide sequences of the respective products of PCR amplification, derived from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction with the identical set of primer pairs used in stage a), and identification of PCR products with essentially identical molecular weights and/or nucleotide sequences;
iii) identification and selection of plants or plant with the desired profile according to the analysis of the markers, in particular plants or plant that contains many of the most favorable alleles at marker loci linked with the corresponding loci QTL.

28. A method of producing plants, which involves the following stages:
(a) crossing two or more parent plants, of which at least one is a plant according to any one of claims 1 to 22 or plant identified by the method according to item 27;
b) screening the progeny from crosses made at the stage a), in the presence of a plant that contains in its genome the entire set of favourable alleles at the corresponding set of loci QTL at least one of the parent plants by:
i. obtaining plant material from a plant-descendant and extracting DNA from this material;
ii. analysis of the DNA sample obtained in stage i)to determine the allelic variants available for a marker loci genetically linked with the corresponding loci QTL, when pomoshnaia markers for p. 25 or 26 when reaction PCR amplification;
iii. identification of marker alleles by determining the molecular weights and/or the nucleotide sequences of PCR products amplification obtained in stage (ii);
c) comparing the identified at stage iii) molecular weights and/or the nucleotide sequences of PCR products amplification with molecular weights and/or the nucleotide sequences of the respective products of PCR amplification derived from inbred lines M/1 (NCIMB 41459) and M/2 (NCIMB 41460) in a PCR reaction with the identical set of primer pairs used in phase ii), and identification of PCR products with essentially identical molecular weights and/or nucleotide sequences;
d) identification and selection of plants or plant with the desired profile according to the analysis of the token.

29. The method according to p, in which stage (a) one of the parent plant has a genetic background, presents the inbred corn line M/1 (NCIMB 41459) or M/2 (NCIMB 41460).

30. The method according to p or 29, in which both parent plants used in the breeding stage (a)are inbred.

31. The method according to item 30, in which the crossing at the stage a) is performed between two parent plants that have a genetic background, presents inbred lines of maize M/1 (NCIMB 41459) and M/2 (NCIMB 41460).

32. The hybrid produced ways the ohms on any of PP-31.

33. The method of applying a set of marker nucleic acids by p. 25 or 26 when selection on markers for the implementation of a set of alleles associated with a corresponding set of loci QTL, idioplasm corn that does not contain a given set of alleles, and these alleles contribute to the phenotypic trait of grain yield by itself or in combination with the moisture content of the grain after the harvest.



 

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2 dwg, 2 ex

FIELD: molecular biology, medicine, biochemistry.

SUBSTANCE: invention proposes a method for assay of mononucleotide changes in the known sequences of nucleic acids. Method involves hybridization with PCR-amplified matrix DNA and the following ligation a tandem on its consisting of tetranucleotide that comprises the diagnosed change and two oligonucleotides of the size 8-10 nucleotides being one of that is immobilized on surface of a solid-phase carrier through a 5'-phosphate linker, and the second oligonucleotide is labeled by 3'-end with biotin label. Then tetranucleotide is hybridized with the matrix DNA chain between two oligonucleotides directly that can be ligated with immobilized oligonucleotide through the 5'-end and with non-immobilized oligonucleotide through the 3'-end. The ligation product is detected by its transformation to enzyme label through the complex with high-affinity enzymatic catalyst followed by development of enzyme label in the presence of chromogenic, luminogenic or fluorogenic substrates. Applying a method provides preparing the simple and highly selective agent used for detection of known changes in gene structure.

EFFECT: improved assay method.

8 cl, 3 dwg, 30 ex

FIELD: genetic engineering, biotechnology, biochemistry, agriculture, food industry, medicine.

SUBSTANCE: invention relates to the transformation of plant with nucleic acid encoding enzyme Δ6-desaturase in C. elegans that results to preparing a plant with enhanced content of gamma-linolenic acid and resistance to cold. Desaturase extracted from the plant can be used for preparing a drug used for treatment of disorder in body associated with deficiency of gamma-linolenic acid in it.

EFFECT: valuable biological properties of genes and desaturases.

36 cl, 9 dwg, 2 ex

FIELD: medicine, hematology.

SUBSTANCE: one should isolate DNA out of peripheral blood lymphocytes due to polymerase chain reaction (PCR) technique of DNA synthesis, carry out genotyping for polymorphism of promoter area of TNF-alpha and TNF-beta gene. While detecting genotype LT*22 being characterized by availability of gene TNF-beta mutation in homozygous state, detect persons predisposed to the development of chronic lympholeukosis, and at certain combinations of genotypes TNF*22/LT*22 or TNF*12/LT*11 in patients with chronic lympholeukosis on should predict an aggressive flow of this disease.

EFFECT: higher accuracy of prediction.

3 ex, 3 tbl

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