Rapeseed pod number major QTL molecular marker and application thereof

A technology of molecular markers and siliques, which is applied in the determination/testing of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve problems such as difficult application, poor performance, and small repeatability, and achieve accurate and rapid screening and accelerated breeding The effect of clear process and position

Active Publication Date: 2015-07-29
INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] At present, there are also some reports on the QTL mapping of rapeseed silique number (Shen Jinxiong et al., 2003; Zhang Shufen et al., 2006; Yi Bin et al., 2006; Gao Bijun et al., 2007; Radoev et al.2008; Shi et al.2009; Wang Feng et al. , 2010; Wu Jianzhong et al., 2010; Sun Meiyu et al., 2013), but usually the detected QTL effect value is small and the repeatability is not good, so it is difficult to apply in rapeseed breeding

Method used

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  • Rapeseed pod number major QTL molecular marker and application thereof
  • Rapeseed pod number major QTL molecular marker and application thereof
  • Rapeseed pod number major QTL molecular marker and application thereof

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Experimental program
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Effect test

Embodiment 1

[0034] Construction and trait determination of silique number segregation populations of rapeseed:

[0035] In the embodiment of the present invention, the F 2 and F 2:3 Separate groups. The silique number phenotypes of the two parents and the two populations were identified after harvesting at the mature stage. The test data of the number of siliques showed that the number of siliques in the two populations was normally distributed, which proved the quantitative genetic characteristics of the number of siliques ( figure 1 ).

Embodiment 2

[0037] Extraction of total DNA from leaves:

[0038] The total DNA of leaves was extracted by the CTAB method, and the specific steps were as follows:

[0039] (1) Take 0.1 g of fresh leaves and grind them, add 700 microliters of extract to grind them, then transfer them to a 1.5 ml centrifuge tube and place them in a constant temperature water bath at 65°C for 60 minutes, during which they mix 2-3 times;

[0040] (2) Add an equal volume of phenol: chloroform: isoamyl alcohol (25:24:1, V / V / V), invert gently to mix well, centrifuge at 12,000rpm for 10 minutes, and gently absorb the supernatant into another 1.5 ml centrifuge tube; add an equal volume of chloroform: isoamyl alcohol (24:1, V / V) and re-extract once;

[0041] (3) Add 1 ml of -20°C pre-cooled absolute ethanol, freeze at -20°C for no more than 30 minutes to allow the DNA to precipitate; / V) Wash with ethanol for 2-3 times, pour off the soaking solution, open the cap of the centrifuge tube and place it in a fume hood...

Embodiment 3

[0044] Primer development and synthesis:

[0045] The SSR primers used by the applicant include two types: synthetic rapeseed public (http: / / www.ukcrop.net / Brassica DB) and self-developed SSR, and STS primers (Wang et al.2012; Huang et al.2013; Shi et al.2014,) PCR amplification was performed on the parental DNA, and the product was electrophoresed in a denaturing polyacrylamide gel. After staining and developing, the size of the band was discriminated, and polymorphic primers were screened. The main software used in the process includes SSRPrimer, BWA and sa mtools; the main reagents include Taq enzyme, dNTP, acrylamide, urea, glacial acetic acid, silver nitrate, etc.

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Abstract

The invention discloses a rapeseed pod number major QTL molecular marker and its application. The marker can be used for marker-assisted selection in pod number character improvement and fine mapping and map-based cloning of QTL site. Primers of the molecular marker are CNU400F:5'-CGAGTTTTTGTGTGTACGTATAGTAAT-3' and CNU400R:5'-CCAAAGTGCGTAAAGGAAGG-3'. Rapeseed Zhongshuang 11 and 73290F3 generation are selected by the use of the marker, and silique number of the screened F3 single plant is higher than that of Zhongshuang 11 by 92.3%. Thus, the marker is utilized for assistant selection so as to greatly enhance selection efficiency of high yield breeding.

Description

technical field [0001] The invention belongs to the fields of rapeseed molecular breeding and biotechnology, and specifically relates to a molecular marker closely linked to the main effect QTL site of Silique number in Brassica napus 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, is the largest source of domestic edible vegetable oil, and plays a very important role in the national edible oil supply security strategy (Wang Hanzhong 2006). In recent years, the annual output of domestic vegetable oil in my country has only been maintained 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 oil su...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 王汉中师家勤杨玉花叶姜刘贵华王新发
Owner INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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