CN107794308B - Specific SNP for identifying wheat grain traits and application thereof - Google Patents
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Abstract
The invention discloses a specific SNP for identifying wheat grain traits and application thereof. The invention provides a method for identifying or assisting in identifying wheat grain traits, which comprises the following steps: detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; the grain character of AA genotype wheat is superior to that of TT genotype wheat; the grain traits are preferably thousand grain weight and/or grain length. The invention develops the SNP locus related to the wheat grain character and designs a primer group based on the KASP technology on the basis of the SNP locus. The primer group provided by the invention is used for identifying the wheat grain characteristics, and has the advantages of rapidness, convenience, accuracy and the like. The method has good application prospect for breeding wheat with different seed properties.
Description
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a specific SNP for identifying wheat grain traits and application thereof.
Background
Wheat is the grain crop with the largest planting area in the world, and in China, the planting area of wheat is second to that of corn and rice and accounts for about 21 percent of the total grain yield. The wheat yield is an important factor directly influencing the living standard of people in China and the national food safety. The improvement of the yield of the wheat and the high and stable yield of the wheat are always main targets pursued by wheat breeders in China for a long time. However, wheat production in China is in a slow development stage at present, and the annual production per unit of wheat is increased by less than 0.8%. With the increase of population, urbanization, land desertification and salinization, the grain planting area is reduced day by day, which is more and more prominent and serious in contradiction with the continuous rigid increase of the grain consumption demand. Therefore, the molecular biology technology is utilized to clone the functional gene related to the wheat yield, and the functional molecular marker is developed, so that an important reference gene resource is provided for the wheat molecular marker-assisted breeding, and the molecular marker-assisted breeding method has important scientific significance and practical application value in the aspects of accelerating the wheat breeding process in China and improving the wheat yield in China.
The grain weight is one of three factors of yield, and the key factors determining the grain weight comprise grain type and grain filling rate. In production and breeding practice, thousand kernel weight is often used as an index for measuring grain size, and is mainly composed of grain type property indexes (such as grain length, grain width, grain thickness and other influence factors) and has a significant positive correlation with yield.
Disclosure of Invention
The invention aims to provide a specific SNP for identifying wheat grain traits and application thereof.
The invention provides a method for identifying or assisting in identifying wheat grain traits, which comprises the following steps: detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; the grain character of AA genotype wheat is superior to that of TT genotype wheat; the grain traits are preferably thousand grain weight and/or grain length.
The invention also provides a method for identifying or assisting in identifying the thousand seed weight of wheat grains, which comprises the following steps: detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; if the wheat is AA genotype and the wheat to be detected is high thousand grain weight wheat; if the genotype is TT, the wheat to be detected is selected as low thousand grain weight wheat; the high thousand grain weight wheat is wheat with the thousand grain weight of more than or equal to 35 g; the low thousand grain weight wheat is wheat with the thousand grain weight of less than 35 g.
The invention also provides a method for identifying or assisting in identifying the grain length of wheat grains, which comprises the following steps: detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; if the wheat is AA genotype and the wheat to be detected is selected as long-grain long wheat; if the genotype is TT, the wheat to be detected is selected as short-grain long wheat; the long-grain length wheat is the wheat with the grain length of more than or equal to 0.65 mm; the short grain length wheat is the wheat with the grain length less than 0.65 mm.
In any of the above methods, the method for detecting whether the genotype based on 2144SNP site in the genomic DNA of wheat to be detected is AA genotype, AT genotype or TT genotype comprises the following steps: and (3) taking the genome DNA of the wheat to be detected as a template, carrying out PCR amplification by adopting a specific primer group, and detecting the PCR amplification product to obtain a genotype result.
The specific primer group is a primer group I or a primer group II;
the primer group I consists of 2144F1, 2144F2 and 2144C;
the primer 2144F1 is (b1) or (b2) as follows:
(b1) a single-stranded DNA molecule shown in sequence 5 of the sequence table;
(b2) DNA molecules obtained by substituting and/or deleting and/or adding one or more nucleotides to the sequence 5 and having the same functions as the sequence 5;
2144F2 is (b3) or (b 4):
(b3) a single-stranded DNA molecule shown in sequence 6 of the sequence table;
(b4) DNA molecules obtained by substituting and/or deleting and/or adding one or more nucleotides to the sequence 6 and having the same functions as the sequence 6;
2144C is (b5) or (b6) as follows:
(b5) a single-stranded DNA molecule shown in sequence 7 of the sequence table;
(b6) and (b) a DNA molecule which is obtained by substituting and/or deleting and/or adding one or more nucleotides in the sequence 7 and has the same function as the sequence 7.
The primer group II consists of TaTPP-F1 and TaTPP-R1;
the TaTPP-F1 is (c1) or (c2) as follows:
(c1) a single-stranded DNA molecule shown in sequence 1 of the sequence table;
(c2) DNA molecules which are obtained by substituting and/or deleting and/or adding one or more nucleotides in the sequence 1 and have the same functions as the sequence 1;
the TaTPP-R1 is (c3) or (c4) as follows:
(c3) a single-stranded DNA molecule shown in a sequence 2 of a sequence table;
(c4) and (b) a DNA molecule which is obtained by substituting and/or deleting and/or adding one or more nucleotides in the sequence 2 and has the same function as the sequence 2.
The primer group I is a primer group based on KASP technology.
The primer group II is a primer group based on the common PCR technology.
When the primer group I is adopted, in a reaction system for PCR amplification, the molar ratio of 2144F1, 2144F2 and 2144C is 12: 12: 30.
when the primer set I is used, the concentration of 2144F1, 2144F2 and 2144C in the reaction system for PCR amplification is 12. mu.M, 2144F2 and 2144C in the reaction system for PCR amplification is 30. mu.M.
When the primer set I is used, the concentration of magnesium ions in a reaction system for PCR amplification is 0.2 mM.
When the primer group I is adopted, the reaction system for PCR amplification specifically comprises: template 2.2. mu.L (DNA content about 100ng), MgCl2mu.L of an aqueous solution 0.04. mu.L, 2 XMaster Mix 2.5. mu.L, 0.056. mu.L of a mixed primer solution (containing 2144F1, 2144F2 and 2144C), and a primer-amplified solution prepared from ddH2Make up to 5. mu.L of O.
The 2 XMaster Mix is collectively called "KASP V4.02 XMaster Mix96/384(Low Rox)", the company LGC of Beijing, catalog No. KBS-1016-017.
When the primer set I is adopted, the reaction procedure of PCR amplification can be specifically as follows: 15min at 94 ℃; 95 ℃ for 20s and a certain temperature for 20s (the temperature of the first circulation is 65 ℃, each circulation is reduced by 1 ℃ compared with the previous circulation), and 10 circulations are carried out; 95 ℃ 20s, 57 ℃ 20s, 30 cycles.
When the primer set I is adopted, PCR amplification is operated on a Quantstudio 7 instrument manufactured by ABI company, and a genotyping result is automatically output. If the result of genotyping is Alle2 (blue dots), the wheat to be tested is AA genotype; if the result of genotyping is Alle1 (red dot), the wheat to be tested is TT genotype; if the genotyping result is Alle1/Alle 2, the wheat to be tested is AT genotype.
The invention also protects the application of the substance for detecting the genotype based on 2144SNP loci in the genome DNA of wheat, which is (d1), (d2), (d3) or (d 4):
(d1) identifying or assisting in identifying the characteristics of the wheat grains; the grain traits are thousand grain weight and/or grain length;
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) identifying or assisting in identifying the grain length of the wheat grains;
(d4) preparing a kit having the use of (d1) or (d2) or (d 3).
The invention also protects specific DNA molecules (molecular markers) as shown in sequence 8 of the sequence table. When W (also denoted by "W") is A, it serves as a molecular marker of high thousand grain weight and/or long grain length. When W (also indicated by "W") is T, the molecular marker is low thousand weight and/or short particle length. The high thousand kernel weight is that the thousand kernel weight of the seeds is more than or equal to 35 g. The low thousand seed weight is that the thousand seed weight of the seeds is less than 35 g. The grain length is more than or equal to 0.65 mm. The short grain length is less than 0.65 mm.
The invention also protects the primer group I or the primer group II.
The invention also protects the application of the primer group I or the primer group II, which is (d1), (d2), (d3) or (d 4):
(d1) identifying or assisting in identifying the characteristics of the wheat grains; the grain traits are thousand grain weight and/or grain length;
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) identifying or assisting in identifying the grain length of the wheat grains;
(d4) preparing a kit having the use of (d1) or (d2) or (d 3).
The invention also provides a kit, which contains the primer group I or the primer group II; the application of the kit is (d1), (d2) or (d 3):
(d1) identifying or assisting in identifying the characteristics of the wheat grains; the grain traits are thousand grain weight and/or grain length;
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) and identifying or assisting in identifying the grain length of the wheat grains.
The invention also protects the application of any one of the methods or the specific DNA molecule or the primer group or the kit in wheat breeding. The breeding aim is to screen wheat with high thousand kernel weight, and wheat with AA genotype based on 2144SNP loci is selected during the breeding. The breeding aim is to screen wheat with low thousand kernel weight, and the breeding is carried out by selecting wheat with TT genotype based on 2144SNP loci. The breeding aim is to screen long-grain wheat, and wheat with the genotype based on 2144SNP loci as the AA genotype is selected during the breeding. The breeding aims to screen wheat with short grain length, and the breeding is carried out by selecting wheat with TT genotype based on 2144SNP loci. The high thousand kernel weight wheat is wheat with kernel thousand kernel weight more than or equal to 35 g. The low thousand grain weight wheat is wheat with the thousand grain weight of less than 35 g. The long-grain length wheat is the wheat with the grain length of more than or equal to 0.65 mm. The short grain length wheat is the wheat with the grain length less than 0.65 mm.
Any 2144SNP site is the 24 th nucleotide from the 5' end of the sequence 8 in the sequence table.
The invention develops the SNP locus related to the wheat grain character and designs a primer group based on the KASP technology on the basis of the SNP locus. The primer group provided by the invention is used for identifying the wheat grain characteristics, and has the advantages of rapidness, convenience, accuracy and the like. The method has good application prospect for breeding wheat with different seed properties.
Drawings
FIG. 1 is partial raw typing data for one of the steps of example 2.
FIG. 2 shows the results of analysis of the thousand kernel weight and the trend of the frequency of A/T allelic variation in cultivars of different generations.
Detailed Description
The following examples are given to facilitate a better understanding of the invention, but do not limit the invention. The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified. The quantitative tests in the following examples, all set up three replicates and the results averaged.
Example 1 discovery of specific SNPs and design of specific primer set
Discovery of specific SNP
Test wheat material: 34 parts of wheat materials (numbered as C1-34, and the specific material information is shown in Table 1) which are distributed in different wheat areas in China and have large grain character difference are selected as the excavation materials of polymorphic sites.
2. Sequence alignment
The following steps of operation are respectively carried out on each wheat material to be tested:
1. extracting the genome DNA of the tested wheat material.
2. And (3) performing PCR amplification by using the genomic DNA extracted in the step (1) as a template and adopting a primer pair consisting of TaTPP-F1 and TaTPP-R1 to obtain a PCR amplification product.
TaTPP-F1 (SEQ ID NO: 1 of the sequence Listing): 5'-CGTGTGGTTGTTTGCGTG-3', respectively;
TaTPP-R1 (SEQ ID NO: 2 of the sequence Listing): 5'-CTAGATATAGGCGAGGGTTATTAC-3' are provided.
3. And (3) taking the PCR amplification product obtained in the step (2) for clone sequencing. 24 clones were tested per wheat material.
4. And (4) carrying out sequence splicing and alignment.
And (3) carrying out A genome sequence evaluation and comparison analysis on 24 clone sequencing results of each wheat material, and finding that two A genome PCR amplification products of different wheat to be tested exist. The two PCR amplification products are 2254bp, the 5 'terminal is identical to TaTPP-F1, the 3' terminal is reverse complementary to TaTPP-R1, the 2121-position and 2168-position nucleotides from the 5 'terminal of one PCR amplification product are shown as sequence 3 in the sequence table, and the 2121-position and 2168-position nucleotides from the 5' terminal of the other PCR amplification product are shown as sequence 4 in the sequence table
Based on the sequence alignment of PCR amplification products of all wheat to be tested, a single SNP is found and named as 2144SNP, which is an A/T polymorphism. 2144SNP, i.e., the 24 th nucleotide from the 5' end of sequence 8 of the sequence Listing.
The genotype of each wheat material tested based on 2144 SNPs is shown in table 1.
5. The tested wheat material is planted in a large net room in the institute of crop science of Chinese academy of agricultural sciences in 2012 10 months in 2012, is subjected to conventional fertigation management, and seeds are harvested in 7 months in 2013, and the thousand kernel weight is measured.
The thousand kernel weight of each wheat material tested is shown in table 1.
TABLE 1
| Numbering | Name (R) | Thousand seed weight | Genotype(s) | Numbering | Name (R) | Thousand seed weight | Genotype(s) |
| C1 | Zhongyou 9507 | 51.7g | AA | C18 | Lumai No. 9 | 26.45g | TT |
| C2 | Zheng Mai 9023 | 44.1g | AA | C19 | Mingxian 169 | 33.2g | TT |
| C3 | Pan 86001-3 | 52.8g | AA | C20 | Anhui No. 3 | 18.29g | TT |
| C4 | Jin Mai No. 8 | 41.3g | AA | C21 | Wheat for emergency | 30.4g | TT |
| C5 | Laizhou 953 | 42.05g | AA | C22 | White winter wheat | 15.75g | TT |
| C6 | Cortex Miscanae Radicis | 44.42g | AA | C23 | Orchid wheat | 28.6g | TT |
| C7 | Three points of cun | 53.66g | AA | C24 | White mango wheat | 29.85g | TT |
| C8 | Purple straw red | 44.35g | AA | C25 | White flower wheat | 24.45g | TT |
| C9 | Red mango | 37.54g | AA | C26 | Chinese spring | 27.35g | TT |
| C10 | Fish and wheat | 44.29g | AA | C27 | Lu Han 328 | 33.7g | TT |
| C11 | Lumai No. 1 | 45.658g | AA | C28 | Nongda 139 | 32.05g | AA |
| C12 | Beijing 15 | 28.55g | TT | C29 | Jingyang 60 | 27.3g | TT |
| C13 | Shijiazhuang 54 | 33.28g | TT | C30 | Nicotiana tabacum 15 | 34.05g | TT |
| C14 | Xuzhou 22 | 51.3g | AA | C31 | White wheat | 24.45g | TT |
| C15 | Wenmai No. 8 | 51.7g | AA | C32 | Twist board | 20.9g | TT |
| C16 | Lankao 906 | 51.7g | AA | C33 | Hongjinmai (Red golden wheat) | 23.4g | TT |
| C17 | Shifeng No. 3 | 34.464g | TT | C34 | All-grass of three months | 28.85g | TT |
Of the 34 wheat tested, 15 were AA genotypes and 19 were TT genotypes based on the 2144SNP genotype. The average thousand grain weight of the wheat to be tested with the AA genotype is 45.91g, and the average thousand grain weight of the wheat to be tested with the TT genotype is 27.54 g.
Taking the thousand kernel weight as a threshold value, wherein the wheat with the thousand kernel weight of more than 35g is the high thousand kernel weight wheat, and the wheat with the thousand kernel weight of less than 35g is the low thousand kernel weight wheat. If the genotype of the wheat to be detected based on 2144SNP is AA type, the wheat to be detected is candidate high thousand grain weight wheat; if the genotype of the wheat to be detected based on 2144SNP is TT type, the wheat to be detected is candidate wheat with low thousand kernel weight. The method identified 93% of the 34 tested wheats with high thousand kernel weight (14/15) and 100% of the 34 tested wheats with low thousand kernel weight (19/19).
Second, design of specific primer set
Designing a primer group based on the KASP technology according to the specific SNP found in the first step as follows:
2144F1 (SEQ ID NO: 5 of the sequence Listing): 5'-GAAGGTGACCAAGTTCATGCTTCACAGACTGCCACATCAGCGGCT-3', respectively;
2144F2 (SEQ ID NO: 6 of the sequence Listing): 5'-GAAGGTCGGAGTCAACGGATTTCACAGACTGCCACATCAGCGGCA-3', respectively;
2144C (SEQ ID NO: 7 of the sequence Listing): 5'-TCTTGATAAATCAGTGCCAGGAG-3', respectively;
application of specific primer group
Respectively carrying out the following steps on each wheat material to be tested in the first step:
1. extracting the genome DNA of the tested wheat material.
2. And (3) performing PCR amplification by using the genomic DNA extracted in the step (1) as a template and adopting the specific primer group designed in the step (II).
Reaction system of PCR amplification: template 2.2. mu.L (DNA content about 100ng), MgCl2mu.L of an aqueous solution 0.04. mu.L, 2 XMaster Mix 2.5. mu.L, 0.056. mu.L of a mixed primer solution (containing 2144F1, 2144F2 and 2144C), and a primer-amplified solution prepared from ddH2O is complemented to5 μ L. In the PCR reaction system, the concentration of magnesium ion was 0.2mM, the concentration of 2144F1 was 12. mu.M, the concentration of 2144F2 was 12. mu.M, and the concentration of 2144C was 30. mu.M. The 2 XMaster Mix is collectively called "KASP V4.02 XMaster Mix96/384(Low Rox)", the company LGC of Beijing, catalog No. KBS-1016-017.
Reaction procedure for PCR amplification: 15min at 94 ℃; 95 ℃ for 20s and a certain temperature for 20s (the temperature of the first circulation is 65 ℃, each circulation is reduced by 1 ℃ compared with the previous circulation), and 10 circulations are carried out; 95 ℃ 20s, 57 ℃ 20s, 30 cycles.
PCR amplification was performed on a Quantstudio 7 instrument manufactured by ABI, and the genotyping results were automatically output. If the result of genotyping is Alle2 (blue dots), the wheat to be tested is AA genotype; if the result of genotyping is Alle1 (red dot), the wheat to be tested is TT genotype; if the genotyping result is Alle1/Alle 2, the wheat to be tested is AT genotype.
And (4) the genotype detection result of each wheat to be tested is consistent with the genotype detection result in the first step.
Example 2 detection of Large samples Using specific primer sets
The individual wheat materials tested are shown in table 2.
First, genotype detection
1. Extracting the genome DNA of the tested wheat material.
2. The genomic DNA extracted in step 1 was used as a template for PCR amplification using the specific primer set designed in step two of example 1.
Reaction system of PCR amplification: template 2.2. mu.L (DNA content about 100ng), MgCl2mu.L of an aqueous solution 0.04. mu.L, 2 XMaster Mix 2.5. mu.L, 0.056. mu.L of a mixed primer solution (containing 2144F1, 2144F2 and 2144C), and a primer-amplified solution prepared from ddH2Make up to 5. mu.L of O. In the PCR reaction system, the concentration of magnesium ion was 0.2mM, the concentration of 2144F1 was 12. mu.M, the concentration of 2144F2 was 12. mu.M, and the concentration of 2144C was 30. mu.M. The 2 XMaster Mix is collectively called "KASP V4.02 XMaster Mix96/384(Low Rox)", the company LGC of Beijing, catalog No. KBS-1016-017.
Reaction procedure for PCR amplification: 15min at 94 ℃; 95 ℃ for 20s and a certain temperature for 20s (the temperature of the first circulation is 65 ℃, each circulation is reduced by 1 ℃ compared with the previous circulation), and 10 circulations are carried out; 95 ℃ 20s, 57 ℃ 20s, 30 cycles.
PCR amplification was performed on a Quantstudio 7 instrument manufactured by ABI, and the genotyping results were automatically output. If the result of genotyping is Alle2 (blue dots), the wheat to be tested is AA genotype; if the result of genotyping is Alle1 (red dot), the wheat to be tested is TT genotype; if the genotyping result is Alle1/Alle 2, the wheat to be tested is AT genotype.
The results of the 2144 SNP-based genotype measurements for each wheat material tested are shown in tables 2, 3 and 4. Part of the raw typing data is shown in FIG. 1.
Second, character detection
2002. In 2005 and 2006, the wheat material to be tested was planted in the south of Henan Luoyang, treated with conventional water and fertilizer, harvested and measured for Thousand Kernel Weight (TKW), grain length (KL) and grain width (KW).
The results for the wheat material tested for the AA genotype are shown in Table 2 (including the results for each wheat tested, as well as the average for all wheat tested for that genotype). The results for the wheat material tested for the AT genotype are shown in Table 3 (including the results for each wheat tested, as well as the average for all wheat tested for that genotype). The results for the test wheat material of the TT genotype are shown in Table 4 (including the results for each test wheat, as well as the average for all test wheat of that genotype). From the trend, the thousand grain weight of the AA genotype wheat is larger than that of the TT genotype wheat, and the grain length of the AA genotype wheat is larger than that of the TT genotype wheat.
The thousand seed weight is more than or equal to 35g, and is defined as the high thousand seed weight; thousand kernel weight < 35g, defined as low thousand kernel weight. The grain length is more than or equal to 0.65mm and is defined as long grain length; grain length < 0.65mm is defined as short grain length. The AA genotype wheat is identified as high thousand grain weight wheat and long grain length wheat, and the accuracy results are shown in Table 2. The TT genotype of wheat was identified as low thousand grain weight wheat and short grain length wheat, and the accuracy results are shown in Table 4.
TABLE 2
TABLE 3
TABLE 4
Third, correlation analysis
The results of the association analysis of the selected varieties in the tested wheat material are shown in Table 5. The results show that: the three-year average thousand kernel weight of the AA genotype wheat to be tested is 41.50g, while the three-year average thousand kernel weight of the TT genotype wheat to be tested is 36.45g, and the difference reaches a very significant level (P < 0.01); in grain length traits, the difference of the wheat material with AA genotype compared with the wheat material with TT genotype reaches a significant or extremely significant level (P <0.05 or P < 0.01). Therefore, compared with the TT genotype, the AA genotype is an excellent grain character genotype.
TABLE 5
Note: p <0.05, P < 0.01.
The results of the association analysis of the local varieties in the tested wheat material are shown in Table 6. The results show that: the three-year average thousand kernel weight of the AA genotype wheat to be tested is 38.9g, while the three-year average thousand kernel weight of the TT genotype wheat to be tested is 31.55g, and the difference reaches a very significant level (P < 0.01); in grain length traits, the difference of the wheat material with AA genotype compared with the wheat material with TT genotype reaches a significant or extremely significant level (P <0.05 or P < 0.01). Therefore, compared with the TT genotype, the AA genotype is an excellent grain character genotype.
TABLE 6
Note: p <0.05, P < 0.01.
Examples 3,
Based on the results of example 2, thousand kernel weight and the frequency of A/T allelic variation of cultivars of different generations were analyzed for trend, and the results are shown in FIG. 2. Along with the advancing of the years, the thousand grain weight of wheat bred varieties in China shows a trend of increasing, and consistent with the change trend, the occurrence frequency of excellent allelic variation A in different varieties in different years also shows a trend of increasing, which shows that modern breeding has a strong selection function on the allelic variation, and the allelic variation is obviously related to the grain weight. Therefore, the marker can be used as a functional marker for improving the wheat grain weight and carrying out wheat high-yield molecular marker assisted breeding.
Sequence listing
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<212>DNA
<213> Artificial sequence
<220>
<223>
<400>7
tcttgataaa tcagtgccag gag 23
<210>8
<211>48
<212>DNA
<213> Artificial sequence
<220>
<221>misc_feature
<222>(24)..(24)
<223>w is a or t
<400>8
tcacagactg ccacatcagc ggcwgctcct ggcactgatt tatcaaga 48
Claims (9)
1. A method for identifying or assisting in identifying wheat grain traits comprises the following steps:
detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; the grain character of AA genotype wheat is superior to that of TT genotype wheat;
the grain properties are preferably embodied as thousand grain weight and/or grain length;
and the 2144SNP site is the 24 th nucleotide from the 5' end of the sequence 8 in the sequence table.
2. A method for identifying or assisting in identifying thousand kernel weight of wheat grains comprises the following steps:
detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; if the wheat is AA genotype and the wheat to be detected is high thousand grain weight wheat; if the genotype is TT, the wheat to be detected is selected as low thousand grain weight wheat;
the high thousand grain weight wheat is wheat with the thousand grain weight of more than or equal to 35 g; the low thousand grain weight wheat is wheat with the thousand grain weight of less than 35 g;
and the 2144SNP site is the 24 th nucleotide from the 5' end of the sequence 8 in the sequence table.
3. A method for identifying or assisting in identifying grain length of wheat grains comprises the following steps:
detecting whether the genotype of a genome DNA of wheat to be detected based on 2144SNP loci is an AA genotype, an AT genotype or a TT genotype; if the wheat is AA genotype and the wheat to be detected is selected as long-grain long wheat; if the genotype is TT, the wheat to be detected is selected as short-grain long wheat;
the long-grain length wheat is the wheat with the grain length of more than or equal to 0.65 mm; the short grain length wheat is the wheat with the grain length less than 0.65 mm;
and the 2144SNP site is the 24 th nucleotide from the 5' end of the sequence 8 in the sequence table.
4. The application of the substance for detecting the genotype based on 2144SNP sites in the genomic DNA of wheat is (d2), (d3) or (d 4):
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) identifying or assisting in identifying the grain length of the wheat grains;
(d4) preparing a kit having the use of (d2) or (d 3);
and the 2144SNP site is the 24 th nucleotide from the 5' end of the sequence 8 in the sequence table.
5. The specific DNA molecule is shown as a sequence 8 in a sequence table.
6. Primer group I consisting of 2144F1, 2144F2 and 2144C;
2144F1 is a single-stranded DNA molecule shown in sequence 5 of the sequence table;
2144F2 is a single-stranded DNA molecule shown in sequence 6 of the sequence table;
2144C is a single-stranded DNA molecule shown in sequence 7 of the sequence table.
7. The use of the primer set according to claim 6, which is (d2), (d3) or (d 4):
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) identifying or assisting in identifying the grain length of the wheat grains;
(d4) preparing a kit having the use of (d2) or (d 3).
8. A kit comprising the primer set of claim 6; the application of the kit is as follows (d2) or (d 3):
(d2) identifying or assisting in identifying the thousand seed weight of the wheat grains;
(d3) and identifying or assisting in identifying the grain length of the wheat grains.
9. Use of the method according to any one of claims 1 to 3 or the specific DNA molecule according to claim 5 or the primer set according to claim 6 or the kit according to claim 8 for wheat breeding.
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| CN107794307B (en) * | 2016-08-29 | 2020-05-12 | 中国农业科学院作物科学研究所 | Specific SNP for identifying wheat grain traits and application thereof |
| CN109022432B (en) * | 2018-09-04 | 2020-06-09 | 中国农业科学院作物科学研究所 | Method for identifying wheat tillering angle character and special primer group thereof |
| CN109468406B (en) * | 2018-12-28 | 2021-12-07 | 中国农业科学院作物科学研究所 | KASP marker related to wheat seedling stage root system configuration and application thereof |
| CN111394506B (en) * | 2020-05-19 | 2023-05-26 | 黑龙江省农业科学院克山分院 | Wheat molecular marker and application thereof in identifying wheat grain weight characteristics |
| CN112266975B (en) * | 2020-11-27 | 2022-07-26 | 山东省农业科学院作物研究所 | Primer group and kit for detecting KASP marker related to POD activity of wheat grains and application of kit |
| EP4111855A1 (en) | 2021-06-29 | 2023-01-04 | INIAV - Instituto Nacional de Invesigação Agrária E Veterinária, I.P. | Snp based panel for mediterranean wheat plant selection and breeding |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102086472A (en) * | 2009-12-08 | 2011-06-08 | 中国农业科学院作物科学研究所 | Gene relevant to thousand seed weight character of wheat, haplotype of gene and application thereof |
| CN104342484A (en) * | 2013-07-23 | 2015-02-11 | 中国农业科学院作物科学研究所 | Molecular marker related with wheat thousand grain weight and applications thereof |
| CN104805081A (en) * | 2015-04-30 | 2015-07-29 | 安徽农业大学 | Wheat grain heavy molecular marker and application thereof |
| CN107794307A (en) * | 2016-08-29 | 2018-03-13 | 中国农业科学院作物科学研究所 | A kind of special SNP for identifying wheat seed character and its application |
-
2016
- 2016-08-29 CN CN201610757883.XA patent/CN107794308B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102086472A (en) * | 2009-12-08 | 2011-06-08 | 中国农业科学院作物科学研究所 | Gene relevant to thousand seed weight character of wheat, haplotype of gene and application thereof |
| CN104342484A (en) * | 2013-07-23 | 2015-02-11 | 中国农业科学院作物科学研究所 | Molecular marker related with wheat thousand grain weight and applications thereof |
| CN104805081A (en) * | 2015-04-30 | 2015-07-29 | 安徽农业大学 | Wheat grain heavy molecular marker and application thereof |
| CN107794307A (en) * | 2016-08-29 | 2018-03-13 | 中国农业科学院作物科学研究所 | A kind of special SNP for identifying wheat seed character and its application |
Non-Patent Citations (3)
| Title |
|---|
| "A Single-Nucleotide Polymorphism of TaGS5 Gene Revealed its Association with Kernel Weight in Chinese Bread Wheat";Shasha Wang等;《Frontiers in Plant Science》;20151223;第6卷;第1-10页 * |
| "Genome-Wide Linkage Mapping of QTL for Yield Components, Plant Height and Yield-Related Physiological Traits in the Chinese Wheat Cross Zhou 8425B/Chinese Spring";Fengmei Gao等;《Frontiers in Plant Science》;20151218;第6卷;第1-17页 * |
| "小麦RILs群体叶绿素含量和千粒重相关分析及QTL定位";刘胜男等;《安徽农业大学学报》;20130712;第40卷(第4期);第570-574页 * |
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