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Molecular markers closely linked with multiple-effect major quantitative trait loci (QTL) of grain weight or silique length of rape, and application thereof

A molecular marker, pleiotropic technology, applied in the direction of recombinant DNA technology, DNA/RNA fragment, microbial determination/inspection, etc., can solve the problems of difficult rapeseed breeding application, poor repeatability, long linkage marker distance, etc. The detection method is convenient and fast, the selection efficiency is improved, and the effect of accurate and rapid screening

Active Publication Date: 2018-01-26
INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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AI Technical Summary

Problems solved by technology

At present, there are some reports on QTL mapping of rapeseed thousand-grain weight (Li et al., 2014), but usually the detected QTL effect value is small, the reproducibility is not good, and the distance from the linkage marker is far away, so it is difficult to apply in rapeseed breeding

Method used

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  • Molecular markers closely linked with multiple-effect major quantitative trait loci (QTL) of grain weight or silique length of rape, and application thereof
  • Molecular markers closely linked with multiple-effect major quantitative trait loci (QTL) of grain weight or silique length of rape, and application thereof
  • Molecular markers closely linked with multiple-effect major quantitative trait loci (QTL) of grain weight or silique length of rape, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Construction of target main effect QTL-NIL

[0029] According to the strategy of fine positioning ( figure 1 ), NIL was constructed for the target main effect QTL site. In this experiment, the parents Zhongshuang 11 and No.73290 were used to cross hybrid F1, and then Zhongshuang 11 was used as the recurrent parent, and the BC5F1 generation QTL-NIL was obtained by continuous backcrossing for 5 generations, and the heterozygous individual plants were selected for further backcrossing . Finally, 80 molecular markers (Yang et al., 2016) other than the target main effect QTL were used for genetic background screening, and the BC5F1 single plants that were heterozygous for the target fragment and had a high background recovery rate were self-crossed to obtain the BC5F2 generation QTL-NIL segregation population. The population was planted in the field as experimental material for fine positioning.

Embodiment 2

[0031] Development of Molecular Markers:

[0032] First, use self-designed primers to compare with the genome sequence of Chinese cabbage or Brassica napus to determine the genomic region corresponding to the target QTL. This genomic region was searched for SSRs using MISA software. Use BWA software to locate the resequencing sequence of 73290 to the reference genome sequence of Zhongshuang11, and use samtools software to find the InDel site of the target QTL interval. Then, use Primer3.0 software to design SSR / InDel primers.

[0033] The genomic DNA of Shuang11, No.73290 and QTL-NIL leaves in the parents was extracted by the magnetic bead method, and the parental DNA was amplified by PCR using self-developed SSR / InDel primers, and the products were electrophoresed in a denaturing polyacrylamide gel , after staining and developing, the size of the bands was discriminated, and polymorphic primers were screened.

[0034] PCR reaction system:

[0035]

[0036] PCR reaction...

Embodiment 3

[0041] Genetic Mapping of Newly Developed Molecular Markers

[0042] Using newly developed markers to screen parental polymorphisms, the present invention has developed five specific co-dominant molecular markers (Ni76, BrSF7-101, BrSF6-2389, BrSF6-2562). To further verify whether these 5 newly developed molecular markers are located in the target major QTL interval, we re-mapped them genetically using the BnaZNF2 population (Shi et al., 2015).

[0043] The genotype is read according to the following principles (such as figure 2 shown), the band type of Shuang11 in the parent is marked as 0 (or A in other embodiments of the present invention), the band type of parent 73290 is marked as 1 (or B in other embodiments of the present invention), and the group The same band type as Zhongshuang 11 is marked as 0 (or A in other embodiments of the present invention), the same band type as 73290 in the population is marked as 1 (or B in other embodiments of the present invention), het...

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Abstract

The invention discloses molecular markers closely linked with multiple-effect major QTL of the grain weight or silique length of rape, and application thereof. The closely-linked molecular markers canbe used for molecular marker-assisted selection of rape and map-based cloning of the QTL. According to the invention, a hybrid F1 and parents are subjected to continuous backcrosses for multiple generations, and a hybrid near-isogenic line is constructed by cooperatively using a molecular marker-assisted selection method; individual plants with high background reversion rate are subjected to selfing so as to obtain a QTL-NIL segregation population; molecular markers are gradually encrypted in a target region so as to realize genotype analysis of the population and searching of crossover individual plants; and thus, the molecular markers BrSF7-101 and BrSF6-2389 are obtained, and the physical distance between the two molecular markers is only 134 kb. The two closely-linked molecular markers are used for genetic typing of linked populations, and it is found that two alleles are greatly different in grain weight and silique length; and thus, the molecular markers can be applied to molecular marker-assisted selection of rape, improve selection efficiency and accelerate breeding progress.

Description

technical field [0001] The invention belongs to the technical field of rapeseed molecular breeding, and in particular relates to a molecular marker closely linked with the pleiotropic main effect QTL of rapeseed grain weight or silique length and its application. Background technique [0002] Rapeseed is one of the most important oil crops in my country, and its planting area and total output account for about one-third of the world (Fu Tingdong, 2010). Rapeseed oil accounts for more than 55% of the oil production of domestic oil crops in my country, and is the largest source of domestic edible vegetable oil. It plays a very important role in the national edible oil supply security strategy (Wang Hanzhong, 2010). In recent years, the annual output of domestic vegetable oil in my country has only remained at about 9-10 million tons, and the self-sufficiency rate is less than 40% and has a further downward trend, which poses a major threat to the security of edible vegetable o...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
Inventor 王汉中师家勤彭伟宋东霁詹杰鹏刘贵华王新发
Owner INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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