CN108707684B - SNP (Single nucleotide polymorphism) marker related to millet flag leaf length and detection primer and application thereof - Google Patents
SNP (Single nucleotide polymorphism) marker related to millet flag leaf length and detection primer and application thereof Download PDFInfo
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- C12Q2600/00—Oligonucleotides characterized by their use
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Abstract
The application discloses an SNP marker related to the leaf length of a millet flag leaf, and a detection primer and application thereof. The SNP marker is positioned in bin markers from 39221591bp to 39480016bp of a second chromosome, and is closely linked with the flag leaf length of millet. The SNP marker related to the millet flag leaf length property can be used for molecular marker assisted breeding of the millet flag leaf length property, the millet flag leaf length can be predicted through SNP marker detection, scientific basis is provided for early identification or screening breeding of the millet flag leaf length property, and important theoretical and practical guiding significance is achieved for accelerating genetic breeding or improvement process of millet varieties. The primer pair for detecting the SNP markers can be used for typing the leaf length of the millet flag and detecting the difference of SNP expression.
Description
Technical Field
The application relates to the technical field of millet breeding, in particular to an SNP marker related to the leaf length of millet flag leaves, and a detection primer and application thereof.
Background
China is the country with the largest cultivation area and the highest yield in the native place of millet and the world, and the yield accounts for about 80 percent of the total amount of the world. Meanwhile, China is also the country with the largest quantity of millet genetic resources and the most abundant diversity.
The flag leaves are the most important photosynthetic organs in the late growth stage of the millet and also are main source organs of carbohydrates of grains, the area indexes such as the length and the width of the flag leaves have very obvious positive correlation with the yield, the flag leaves are important factors influencing the yield of the millet, genes related to yield characters such as the length and the width of the flag leaves of the millet and position functions of the genes are researched and controlled, and the method has important significance for guiding genetic breeding of the millet.
Single Nucleotide Polymorphism (SNP) is a DNA sequence polymorphism caused by variation of a single nucleotide base at the genome level, and is large in number, widely distributed, and good in genetic stability in the genome. With the development of gene sequencing technology and the reduction of cost, the same gene or gene fragment of different individuals is directly sequenced and compared with the sequence, so that whether the base has variation or not can be determined. Therefore, SNP detection is beneficial to genotyping and is suitable for rapidly and massively screening the relation between unknown or known SNP and certain genetic traits.
At present, QTL positioning and SNP marker research related to the quantitative traits of millet mainly focuses on the development and utilization of SSR markers, and the like, but the development and application research of SNP molecular markers for detecting polymorphism by utilizing degenerate genome sequencing of a population inbred line has not been reported. Therefore, the development of the quantitative character SNP molecular marker of the millet is developed, and an auxiliary breeding system is established, so that the method has important significance for improving the yield of the millet and saving the breeding cost.
Disclosure of Invention
The purpose of the application is to provide a novel SNP marker related to the leaf length of millet flag leaves, and a detection primer and application thereof.
In order to achieve the purpose, the following technical scheme is adopted in the application:
one aspect of the present application discloses an SNP marker associated with the shape of the leaf length of a millet flag, the SNP marker is located within the bin markers of the 39221591bp to the 39480016bp of the second chromosome, and the SNP marker is closely linked with the leaf length of the millet flag.
It should be noted that, through research, a sequence closely linked to the length of the flag leaf of millet is discovered, and the SNP marker in the sequence directly affects the trait of the length of the flag leaf of millet, i.e. the bin marker from 39221591bp to 39480016bp of the second chromosome.
Further research shows that the bases of the bin markers from 39221591bp to 39480016bp of the second chromosome, especially SNP markers, are C or G and are closely linked with the leaf length of the millet flag.
In one embodiment of the present application, one of the SNP markers, that is, the FLL-HAI-02-061 site is located at 39309061bp of the sequence of the second chromosome, the FLL-HAI-02-061 site is located in the sequence shown in Seq ID No.1, and the FLL-HAI-02-061 site is 37 th base from the 5' end of the sequence shown in Seq ID No. 1.
It should be noted that bin markers from 39221591bp to 39480016bp of the second chromosome are closely linked with the length of the flag leaf of millet, and may contain a plurality of SNP markers, the FLL-HAI-02-061 site is only the SNP marker related to the shape of the long leaf of millet confirmed in one implementation mode of the application, and other SNP markers in the bin markers are not excluded under the guidance of the application.
Preferably, the SNP marker related to the long characteristic of the millet flag leaf of the application utilizes a composite interval mapping method for the SNP marker, the total significance level is 5%, the LOD value detected by QTL is 3.4857, and the phenotypic variation interpretation rate is 3.08%.
The other side of the application discloses a primer pair for detecting the SNP marker of the application, wherein an upstream primer of the primer pair is a sequence shown by Seq ID No.2, and a downstream primer of the primer pair is a sequence shown by Seq ID No. 3;
Seq ID No.2：5’-GCCGTCTGAATGATGGTGC-3’
Seq ID No.3：5’-AGGTGGACTGCGAGCTGAA-3’。
the other side of the application discloses a kit for detecting the SNP marker of the application, and the kit comprises the primer pair of the application.
Preferably, the kit of the present application further comprises reagents for PCR amplification.
The application also discloses a method for detecting the SNP marker, which comprises the steps of carrying out PCR amplification on the genomic DNA of the millet to be detected by adopting the primer pair or the kit, and obtaining the base condition of the SNP marker according to the sequencing result of the PCR amplification product.
The application also discloses a method for predicting the length of the millet flag leaves, which comprises the steps of carrying out PCR amplification on the millet genome DNA of a prediction object by adopting the primer pair or the kit, obtaining the base condition of the SNP marker of the application according to the sequencing result of the PCR amplification product, and predicting the length of the millet flag leaves according to the base condition of the SNP marker.
The application further discloses application of the SNP marker in millet flag leaf length prediction, early millet flag leaf length identification or millet molecule auxiliary screening breeding.
The application further discloses application of the primer pair or the kit in millet flag leaf length prediction, early millet flag leaf length identification or millet molecule auxiliary screening breeding.
Due to the adoption of the technical scheme, the beneficial effects of the application are as follows:
the SNP marker related to the millet flag leaf length property can be used for molecular marker assisted breeding of the millet flag leaf length property, the millet flag leaf length can be predicted through SNP marker detection, scientific basis is provided for early identification or screening breeding of the millet flag leaf length property, and important theoretical and practical guiding significance is achieved for accelerating genetic breeding or improvement process of millet varieties. The primer pair for detecting the SNP markers can be used for typing the leaf length of the millet flag and detecting the difference of SNP expression.
Drawings
FIG. 1 is an electrophoretogram of parental genomic DNA of millet in the present example, wherein the first lane is lane M, which represents DL2000marker, the second lane, which is the lane marked with 1, represents genomic DNA of paternal "Ji Zhang Gu No." and the third lane, which is the lane marked with 2, represents genomic DNA of maternal "A2";
FIG. 2 is an electrophoresis diagram of PCR products obtained by amplifying parental genomic DNA of foxtail millet by using a primer pair of 12_2F and 12_2R in the present example, wherein the first lane is a lane marked with A12-2, which shows the PCR product using maternal "A2" genomic DNA as a template, the second lane is a lane marked with 312-2, which shows the PCR product using paternal "Ji Zhang Gu No. I" genomic DNA as a template, and the third lane is an M lane, which shows DL2000 marker;
FIG. 3 is a diagram showing the alignment results of the sequences of FLL-HAI-02-061 loci in the female and male SNP markers in the present example, the FLL-HAI-02-061 loci are in the bin markers of 39221591bp to 39480016bp of the second chromosome, the female parent base is C, and the male parent base is G.
Detailed Description
The application provides SNP markers of bin marker regions related to the length of the leaf of the Valeriana officinalis L, detection primers and application thereof. The SNP marker FLL-HAI-02-061 closely linked to the length of the millet flag leaf is located in the bin marker of 39221591bp to 39480016bp of the second chromosome.
In this application, Bin means Bin mapping (i.e., Bin map), and the minimum unit (Recombination Bin) constituting a Recombination event is obtained using all Recombination site information in the mapped population. The obtained bin is used as a marker for subsequent linkage map construction and QTL positioning.
In the present application, a genetic map (genetic map or linking map) refers to a linear arrangement of relative positions between markers constructed on the basis of the recombination rate between genetic markers. Sequencing parent and filial generation populations of the mapping population based on a high-throughput resequencing technology, and taking a certain number of continuous SNPs as a basis for judging recombination sites of the filial generation to obtain a whole genome physical recombination map of each filial generation. Using all Recombination site information in the mapped population, the minimum unit (Recombination Bin) and Bin map (i.e., Bin map) that constitute the Recombination event are obtained and the resulting bins are used as markers for subsequent linkage map construction and QTL mapping.
The application provides male parent 'Ji Zhang Gu No. I' and female parent 'A2' materials of millet, F1 generation materials of hybridization of the male parent and the female parent, and 441 parts of self-bred 13 generation F2 group materials. Planting all the materials in the Hainan area, and recording and arranging character data such as flag leaf length and the like. And after carrying out degenerate genome re-sequencing on each material, completing comparison and reference genome splicing to obtain the SNP polymorphic marker. And obtaining related SNP markers through flag leaf length data, and verifying the SNP markers through clone sequencing.
The present application is described in further detail below with reference to specific embodiments and the attached drawings. The following examples are intended to be illustrative of the present application only and should not be construed as limiting the present application.
Example high throughput sequencing and SNP marker information analysis
This example uses the male parent "Ji Zhang Gu I" and female parent "A2" material, the F1 generation material of the hybridization of the two, and the F2 group 441 parts material of the selfing 13 generation, i.e. recombination selfing line (RILs). Planting all the materials in the Hainan area, and recording and arranging character data such as flag leaf length and the like. And after carrying out degenerate genome re-sequencing on each material, completing comparison and reference genome splicing to obtain the SNP marker. And (3) obtaining related SNP markers by analyzing the flag leaf length data, and verifying the SNP markers by clone sequencing.
In this example, genomic DNA of each sample was extracted by RADSeq method, and a total of 444 samples including 441 RILs, 2 parents and 1F 1 individuals were obtained. Performing quality detection on the extracted genome DNA, performing enzyme digestion on the qualified DNA, recovering DNA fragments through electrophoresis, adding a joint for cluster preparation, and finally performing machine sequencing.
The specific steps of this example are as follows:
(1) weighing 1.0g of fresh leaves, shearing, putting into a mortar, grinding with liquid nitrogen, adding 3mL of 1.5 xCTAB, grinding into homogenate, transferring into a 15mL centrifuge tube, adding 1mL of 1.5 xCTAB into the mortar for washing, transferring into the centrifuge tube, uniformly mixing, and slowly shaking at intervals in a water bath at 65 ℃ for 30 min.
Wherein 1L of a 1.5 × CTAB formulation comprises: 15g of CTAB, 1mol/L of Tris.Cl 75mL with the pH value of 8.0, 0.5mol/L EDTA 30mL and 61.4g of NaCl, and adding deionized water to fix the volume to 1L; before use, about 2mL of mercaptoethanol was added to a final concentration of 0.2%.
(2) After the water bath is finished, cooling to room temperature, adding equal volume of chloroform/isoamylol, and gently mixing uniformly until the subnatant turns into dark green; wherein the volume ratio of chloroform to isoamyl alcohol is 24: 1.
(3) Centrifuging at 4200rpm for 10min, transferring the upper aqueous phase to a new 15mL centrifuge tube, adding 2 times volume of precooled absolute ethanol, mixing and standing for 5 min. The DNA was precipitated by standing at-20 ℃ for 30 min.
(4) Centrifuging at 4200rpm for 10min, discarding the supernatant, adding 1mL of 75% ethanol to wash the precipitate 1 time, inverting the centrifuge tube to dry the DNA, and adding 50. mu.L of TE to dissolve the DNA.
(5) The concentration of DNA was measured and adjusted to 20 ng/. mu.L with water.
(6) And digesting by adopting PstI enzyme to break the genome DNA, wherein the digestion reaction system of the PstI enzyme is 30 mu L and comprises the following steps: genomic DNA 20. mu. L, PstI enzyme 1. mu.L, 10 XPstI buffer 3. mu. L, Nuclease-free water 6. mu.L. After the enzyme digestion reaction system is prepared, incubation is carried out for at least 1 hour at the constant temperature of 37 ℃.
(7) And (3) performing a joint reaction, namely adding a sequencing joint to the broken genomic DNA fragment, wherein the joint reaction system is 40 mu L and comprises the following steps: 30 μ L of PstI digestion product, 1 μ L of sequencing linker, 10 Xligation buffer 1 μ L, T4DNA ligase 2 μ L, rATP 0.4.4 μ L, ddH2O 5.6μL。
(8) mu.L of each reaction product was taken and added to a new centrifuge tube, one set of 12 samples each, and the total volume of each set was 12. mu.L.
(9) Electrophoresis was performed on a 3% recovery gel for 1h, and the 300-and 700 bp-sized fragment was excised after EB staining. Gel purification was performed with QIAquick Kit and the recovered product was dissolved in 30. mu.L of EB solution.
(10) Carrying out PCR reaction, carrying out PCR amplification on the cut and recovered fragments by adopting a universal primer of a sequencing joint, wherein the PCR reaction system is 25 mu L and comprises the following steps: 10 XBuffer 2.5 uL, 25mM dNTPs 0.25 uL, 5U/. mu.L Hi-Fi enzyme 0.25 uL, 10 uM sequencing adaptor forward primer 0.5 uL, 10 uM sequencing adaptor reverse primer 0.5 uL, gel cutting recovery product 20 uL, sterile water 1 uL.
The PCR reaction conditions were 94 ℃ pre-denaturation for 5min, followed by 10 cycles: denaturation at 94 ℃ for 30 seconds, annealing at 60 ℃ for 30 seconds, extension at 72 ℃ for 40 seconds, and extension at 72 ℃ for 3min after the cycle is finished.
(11) And (5) magnetic bead purification to complete library construction.
The specific purification method comprises the following steps: first, 1.2-fold volume of magnetic beads was added to the PCR product, and the mixture was allowed to stand for 10 min. Then, the mixture was adsorbed on a magnetic frame, and the supernatant was removed. Then, 500. mu.L of 70% ethanol was added to wash twice. After drying to dryness on a dry heat instrument, 15 μ L of EB solution was added and dissolved for 5 min. Finally, the supernatant was transferred to a 1.5mL centrifuge tube by adsorption on a magnetic rack.
(12) And (5) performing on-machine sequencing after the detection reaches the on-machine standard.
As a result: enzyme digestion, library building and sequencing are carried out on 444 samples, 75.99Gb raw data are obtained, and 171.54Mb of each individual is averaged. The sequencing sequences were aligned to the reference genome with an average alignment of 86.326%, an average coverage of 8.226%, and an average sequencing depth of 3.655 ×.
Wherein, 444 samples comprise 441 RILs, 2 parents and 1F 1 individuals; the reference genome in this example, Yugu genome, can be obtained from the following website:
https://www.ncbi.nlm.nih.gov/genome/?term＝Setaria+italica+(foxtail+millet)。
33771 SNP markers with polymorphism between parents are obtained by sequencing and information analysis. Selecting a plurality of SNPs as a window according to a window sliding method, sliding one SNP every time to determine the genotype of each window and the crossover site of each individual, and generating a bin genotype and a bin map. According to the bin genotype data, MSTMap software is used for constructing a genetic map, 2022 bins are positioned on 9 chromosomes, and then the genetic map data are introduced into MapChart software to integrate a genome genetic linkage map.
And carrying out QTL analysis on the long trait phenotype of the flag leaves of the millet by using the constructed high-density genetic map of the millet and adopting composite interval mapping analysis (CIM). QTL detection adopts 5% of total significance level, and according to 500 times of arrangement test results, the critical LOD value of flag leaf length trait phenotype data QTL analysis is determined, and prediction bin interval and SNP locus information related to the flag leaf length are obtained through analysis.
The results showed that the SNP marker associated with the long character of the millet flag leaf is located in 241 centimorgans (cM), 205 th bin to 206 th bin (i.e. chr2_ bin205-chr2_ bin206) of the second chromosome of the genetic linkage map, namely, the bin marker of the 39221591bp to 39480016bp of the second chromosome; wherein, the FLL-HAI-02-061 locus is positioned at the 39309061bp position of the sequence of the second chromosome; the bases of these SNP markers related to the leaf length of millet are all C or G. By using a composite interval mapping method, the total significance level of 5% is adopted, the LOD value detected by QTL is 3.4857, the phenotypic variation interpretation rate is 3.08%, and the Additive effect value (A) is-1.3808.
EXAMPLE two SNP marker validation
And (3) comparing the reference genome according to the predicted bin marker and the SNP locus to obtain a gene sequence of a related region, selecting about 300bp before and after the SNP locus, designing and developing SNP marker primers, and performing PCR amplification by taking male parent and female parent material DNAs as templates. Selecting the amplification products with normal amplification of the primers and the composite predicted size of the PCR amplification products to carry out gel cutting recovery, sequencing, and selecting the marker primers with the SNP locus difference of the male parent material amplification gene sequences and the female parent material amplification gene sequences. And (3) carrying out SNP marker primer PCR amplification and result sequencing by taking F1 generation and F2 generation materials as templates, and selecting a marker primer which accords with genotyping and is related to phenotypic traits.
Firstly, according to the sequencing result after PCR product recovery, the marker of the difference of the SNP loci of the amplified gene sequences of the male parent material and the female parent material is selected. The method comprises the following specific steps:
(1) according to the steps (1) to (4) in example one, male and female genomic DNAs were extracted by CTAB method, respectively.
(2) The extracted male parent and female parent genomic DNAs were detected by 0.8% agarose gel electrophoresis, and the detection results are shown in FIG. 1, wherein the second lane, lane labeled 1, represents the male parent "Ji Zhang Gu No.' genomic DNA, and the third lane, lane labeled 2, represents the female parent" A2 "genomic DNA. Therefore, the male parent and the female parent genome DNA can be successfully extracted.
(3) Storing the extracted male parent and female parent genome DNA at-20 ℃ for later use.
(4) And respectively taking the extracted male parent genomic DNA and the extracted female parent genomic DNA as templates, and amplifying the genomic DNA by adopting a primer pair 12_2F and a primer pair 12_ 2R. The upstream primer 12_2F of the primer pair is a sequence shown in Seq ID No.2, and the downstream primer 12_2R is a sequence shown in Seq ID No. 3;
Seq ID No.2：5’-GCCGTCTGAATGATGGTGC-3’
Seq ID No.3：5’-AGGTGGACTGCGAGCTGAA-3’。
PCR reaction 25. mu.L, comprising: 10 XBuffer 2.5 uL, 25mM dNTPs 0.15 uL, 5U/. mu.L Taq enzyme 0.15 uL, 10 uM upstream primer 0.5 uL, 10 uM downstream primer 0.5 uL, genomic DNA 1.0 uL, sterile water 20.2 uL.
The PCR reaction conditions were 94 ℃ pre-denaturation for 5min, followed by 35 cycles: denaturation at 94 ℃ for 30 seconds, annealing at 60 ℃ for 30 seconds, extension at 72 ℃ for 40 seconds, and extension at 72 ℃ for 3min after the cycle is finished.
The PCR amplification product was purified and stored at 4 ℃. A portion of each PCR amplification product was detected by 1% agarose gel electrophoresis, and the results are shown in FIG. 2. FIG. 2 is an electrophoretogram of PCR products amplified by the primer pair 12_2F and 12_ 2R. The results in FIG. 2 show that the designed primer pair can amplify the genomic DNA of the male parent and the female parent to obtain target fragments which are in accordance with the expectation.
In this example, the amplified products of the male and female parent genomic DNAs amplified by the primer pairs 12_2F and 12_2R were sequenced, respectively, the sequenced sequences of the male parent amplified products amplified by the primers 12_2F and 12_2R were the sequence shown in Seq ID No.1, and the FLL-HAI-02-061 site was the 37 th base from the 5' end in the sequence shown in Seq ID No. 1. The sequencing sequence alignment results of the male parent amplification product and the female parent amplification product are shown in fig. 3, wherein the base of the background marked with gray represents the base of the FLL-HAI-02-061 locus, the base of the female parent is C, and the base of the male parent is G.
Then, screening according to the leaf length data of the sample: the leaf length of the male parent flag leaf is 31cm, and the leaf length of the female parent flag leaf is 17 cm. Among 441 samples, the minimum value was 6cm, and the maximum value was 42 cm; the number of samples which are the same as the maternal SNP in 50 samples with the smallest length of the flag leaves is more than 80 percent; the number of samples identical to the paternal SNP in the 50 samples with the largest length of flag leaves is more than 70%.
The foregoing is a more detailed description of the present application in connection with specific embodiments thereof, and it is not intended that the present application be limited to the specific embodiments thereof. It will be apparent to those skilled in the art from this disclosure that many more simple derivations or substitutions can be made without departing from the spirit of the disclosure.
SEQUENCE LISTING
<110> Zhang Kouzhou agricultural science institute (alpine crop research institute in Hebei province)
<120> SNP marker related to millet flag leaf length, detection primer and application thereof
<130> 17I25627
<160> 3
<170> PatentIn version 3.3
<210> 1
<211> 298
<212> DNA
<213> sequence of FLL-HAI-02-061 site
<400> 1
tggtgccacc gtgccagcac cggcgcagca gcgcctgggc gctaacctcc aactcactct 60
cctctccttg tctcctcgat ctccctctct ggccggtctc ccctcaacgc cgtcgcaccc 120
ccgtcaatta agcactccga ttgcttgccg ccgggacgga gtccgccggc gctgtctgac 180
tatatacgac ggcgcccccg cccctccggt gcactcccca gtatcgccta cggcgtcttc 240
gccacctctg gctctttgtg ctggatcatg tagctagttc tcgttgatgt cttcagct 298
<210> 2
<211> 19
<212> DNA
<213> Artificial sequence
<400> 2
gccgtctgaa tgatggtgc 19
<210> 3
<211> 19
<212> DNA
<213> Artificial sequence
<400> 3
aggtggactg cgagctgaa 19
Claims (4)
1. An application of SNP marker in molecular assisted screening and breeding in predicting millet flag leaf length, identifying early millet flag leaf length or determining millet flag leaf length character;
the SNP marker is a sequence shown in Seq ID No.1 and has a SNP locus, wherein the 37 th base from the 5' end of the sequence shown in Seq ID No.1 is C or G; the SNP locus is positioned in bin markers from 39221591bp to 39480016bp of a second chromosome and is tightly linked with the flag leaf length of millet; the SNP site is located at the 39309061bp position of the sequence of the second chromosome.
2. Use according to claim 1, characterized in that: for SNP loci, a composite interval mapping method is utilized, the total significance level of 5% is adopted, the LOD value detected by QTL is 3.4857, and the phenotypic variation interpretation rate is 3.08%.
3. The application of a primer pair in molecular assisted screening breeding in predicting the length of a millet flag leaf, identifying the early stage of the length of the millet flag leaf or identifying the long character of the millet flag leaf;
the upstream primer of the primer pair is a sequence shown by Seq ID No.2, and the downstream primer is a sequence shown by Seq ID No. 3;
Seq ID No.2：5’- GCCGTCTGAATGATGGTGC -3’；
Seq ID No.3：5’- AGGTGGACTGCGAGCTGAA -3’；
the primer pair is used for detecting an SNP marker, the SNP marker is a sequence shown by Seq ID No.1 and has an SNP locus, wherein the 37 th base from the 5' end of the sequence shown by Seq ID No.1 is C or G; the SNP locus is positioned in bin markers from 39221591bp to 39480016bp of a second chromosome and is tightly linked with the flag leaf length of millet; the SNP site is located at the 39309061bp position of the sequence of the second chromosome.
4. Use according to claim 3, characterized in that: for SNP loci, a composite interval mapping method is utilized, the total significance level of 5% is adopted, the LOD value detected by QTL is 3.4857, and the phenotypic variation interpretation rate is 3.08%.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104488488A (en) * | 2014-11-23 | 2015-04-08 | 中国科学院遗传与发育生物学研究所 | Method for judging and selecting dim light tolerance |
| CN107090494A (en) * | 2017-03-24 | 2017-08-25 | 张家口市农业科学院 | The molecular labeling related to millet code grain number character and its detection primer and application |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104488488A (en) * | 2014-11-23 | 2015-04-08 | 中国科学院遗传与发育生物学研究所 | Method for judging and selecting dim light tolerance |
| CN107090494A (en) * | 2017-03-24 | 2017-08-25 | 张家口市农业科学院 | The molecular labeling related to millet code grain number character and its detection primer and application |
Non-Patent Citations (1)
| Title |
|---|
| Identification of QTLs for 14 Agronomically Important Traits in Setaria italica Based on SNPs Generated from High-Throughput Sequencing;Kai Zhang等;《G3: GENES GENOMES GENETICS》;20170331;第7卷(第5期);摘要，表2，附表S1、S5 * |
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