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Strong winterness brassia campestris L. soluble protein content molecular markers and QTL location

A cabbage-type winter rapeseed, protein content technology, applied in the fields of molecular biology and genetic breeding, can solve the problem that the research on molecular markers and QTL mapping has not yet been reported, and achieves the advantages of speeding up the breeding process, improving the screening efficiency and saving the production cost. Effect

Active Publication Date: 2015-10-21
GANSU AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, there have been many studies on the content and variation of soluble protein in Chinese cabbage-type winter rapeseed, but there have been no reports on its molecular markers and QTL mapping.

Method used

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  • Strong winterness brassia campestris L. soluble protein content molecular markers and QTL location
  • Strong winterness brassia campestris L. soluble protein content molecular markers and QTL location
  • Strong winterness brassia campestris L. soluble protein content molecular markers and QTL location

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: Construction of isolated populations of strong winter Chinese cabbage-type winter rapeseed and determination of soluble protein content.

[0028] The specific construction of the isolated population used in this embodiment is as follows:

[0029] (1) In August 2010, multi-generation bagging and self-crossing of the parent rapeseed 'Longyou 7' with strong winter resistance and strong winter resistance and rapeseed 'Longyou 9' with strong winter resistance were used to configure the hybrid combination, harvested in May 2011 F1 generation seeds.

[0030] (2) F1 seeds were sown in August 2011, and bagged and self-crossed in April 2012 to obtain F2 generation seeds.

[0031] The F2 generation seeds were sown in August 2012. At the seedling stage, 103 plants were randomly selected to be listed and marked, and fresh young leaves were collected in early November, brought back to the laboratory in an ice box and stored in a -70°C ultra-low temperature refrigerator. ...

Embodiment 2

[0032] Example 2: Extraction of total DNA from leaves of parents and F2 generation segregation populations.

[0033] Total DNA was extracted from the leaves by the CTAB method, and the specific steps were as follows:

[0034] (1) Pick healthy young leaves as materials for extracting rapeseed genomic DNA, wash them with distilled water, and dry them with paper.

[0035] (2) Take about 0.5g of fresh leaves and put them in a mortar, add liquid nitrogen, grind them quickly until they are whitish powdery, and put them into a 2ml centrifuge tube immediately.

[0036] (3) Add 700ul of preheated 2×CTAB extraction buffer to infiltrate the powder and invert the centrifuge tube to fully disperse the powder. Place in a water bath at 65°C for 60 minutes, during which time gently mix 2-3 times.

[0037] (4) Take out the centrifuge tube, cool to room temperature, add 700ul of chloroform / isoamyl alcohol (24 / 1), gently invert to mix, and let stand for 10 min.

[0038] (5) Centrifuge at 1300...

Embodiment 3

[0046] Example 3: Source, synthesis and polymorphism screening of SSR and InDel primers.

[0047] (1) 315 pairs were uniformly selected from the rapeseed microsatellite primer sequences published on www.UK crop.net and all primers on the 10 chromosomes published on the Brassica Database website http: / / brassicadb.org / . Among them, there are 51 pairs of SSR primers and 264 pairs of InDel primers. Primers were synthesized by Shanghai Bioengineering Technology Co., Ltd.

[0048] (2) Randomly select 6 strains of DNA from each parent and mix them in equal amounts, and use them as templates for screening primers.

[0049] (3) PCR reaction system

[0050] The PCR reaction system is 10ul:

[0051] Mix Taq Enzyme 6ul

[0052] Upper primer 0.5ul

[0053] Lower primer 0.5ul

[0054] ddH2O 2ul

[0055] Template: 1ul

[0056] (4) PCR amplification program: pre-denaturation at 94°C for 4 minutes; denaturation at 94°C for 50s, annealing at 55-60°C for 50s (annealing temperature decrease...

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Abstract

The invention discloses strong winterness brassia campestris L. soluble protein content molecule markers and QTL loci. The screening steps are that (1), multi-generation bagging selfing parent strong winterness superstrong cold resistant winter rape 'the 7th Longyou' and strong cold resistant winter rape 'the 9th Longyou' are utilized for hybridization to construct a generation F2 segregation population; (2), a CTAB method is utilized for extracting leaf DNA of the generation F2 segregation population and the two parents; (3), SSR and InDel primers are utilized for performing polymorphism primer selection on the DNA of the two parents; (4), genetype data are obtained through the generation F2 segretation population to construct a genetic linkage map and QTL analysis; and (5), two strong winterness brassia campestris L. soluble protein content QTL loci are obtained and named as Qsol.gsau-8A; and two molecule markers linked with the strong winterness brassia campestris L. soluble protein content QTL loci are BrID10839 and Ra2-E12.

Description

technical field [0001] The invention belongs to the technical field of molecular biology and genetic breeding, and specifically relates to a QTL site for soluble protein content of strong winter Chinese cabbage type winter rapeseed, and also relates to a molecule linked to the QTL site for soluble protein content of strong winter Chinese cabbage type winter rape mark. Background technique [0002] Cabbage-type rape has a long history of cultivation in my country, and has the characteristics of cold resistance, early maturity, and barren resistance, and is one of the important oil crops in my country. But Chinese cabbage-type rapeseed is mainly spring-type, and there is a shortage of germplasm of strong-winter-type rapeseed, especially the varieties of strong-winter-type rape that are suitable for planting in the northern arid and cold regions. The climate in northern my country is very cold, and winter rapeseed has excellent cold resistance to survive the winter safely. The...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 孙万仓方彦杨刚刘自刚曾秀存武军艳李学才孔德晶王凯音马骊
Owner GANSU AGRI UNIV
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