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Molecular marker, primer pair and molecular marking method used for identifying flowering time of brassica oleracea and application of molecular marker, primer pair and molecular marking method

A technology of molecular markers and cabbage, applied in the field of molecular biology, can solve the problems of heavy workload, long cycle, cumbersome operation procedures, etc., and achieve the effect of overcoming the long time cycle

Active Publication Date: 2017-09-22
SOUTHWEST UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there are mainly two methods for screening breeding materials. One is the traditional breeding method. This method requires the selection and identification of bolting resistance traits across the year. The cycle is long, the operation procedures are cumbersome, and the workload is heavy. In addition to being affected by genetic factors, it is also closely related to environmental factors, so relying on phenotypic identification has greater inaccuracy; another method is the molecular marker-assisted method developed in recent years, which combines molecular marker technology with traditional breeding. Combining, using stable and reliable DNA molecular markers to replace environmental sensitive phenotypic markers to screen breeding materials

Method used

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  • Molecular marker, primer pair and molecular marking method used for identifying flowering time of brassica oleracea and application of molecular marker, primer pair and molecular marking method
  • Molecular marker, primer pair and molecular marking method used for identifying flowering time of brassica oleracea and application of molecular marker, primer pair and molecular marking method
  • Molecular marker, primer pair and molecular marking method used for identifying flowering time of brassica oleracea and application of molecular marker, primer pair and molecular marking method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Statistics of early flowering cabbage and late flowering cabbage

[0048] The field flowering time of different cabbage was investigated, and 26 early-flowering cabbage and 24 late-flowering cabbage were screened out. Statistics of the flowering time of early-flowering and late-flowering cabbage for three consecutive years are as follows: in 2015, the average flowering time of early-flowering cabbage was 193.23±8.10 days, and the average flowering time of late-flowering cabbage was 203.5±9.57 days. The flowering time of flowering cabbage is about 10 days later than that of early flowering cabbage; in 2016, the average flowering time of early flowering cabbage was 199.77±8.32 days, and the average flowering time of late flowering cabbage was 215.96±7.77 days. Early flowering cabbage delayed flowering time for about 16 days; in 2017, the average flowering time of early flowering cabbage was 198.29 ± 11.75 days, and the average flowering time of late flowering ca...

Embodiment 2

[0049] Example 2 Extraction of cabbage genomic DNA

[0050] (1) Get 50 parts of young leaves of cabbage cabbage screened in Example 1, store them in 1.5mL centrifuge tubes, and grind them into pulverized shapes in liquid nitrogen;

[0051] (2) Add a mixed solution prepared by mixing 700 μl CTAB extract and 14 μl β-mercaptoethanol at 65°C into each centrifuge tube containing the sample, bath at 65°C for 45 minutes, and mix 3-4 times in between;

[0052] (3) add the chloroform / isoamyl alcohol mixed solution with an equal volume ratio of the mixed solution obtained in step (2) to 24:1 into each centrifuge tube containing the sample, shake gently, mix evenly, emulsify for 10 min, and centrifuge at 10000 rpm 10min, take the supernatant into a new centrifuge tube;

[0053] (4) adding the chloroform / isoamyl alcohol mixed solution whose volume ratio equal to the volume ratio of the supernatant obtained in step (3) is 24:1 to the new centrifuge tube containing the supernatant obtained...

Embodiment 3

[0057] Example 3 Anchoring of BoFLC2 gene differential fragments

[0058] (1) According to the BoFLC2 reference genome sequence, use Primer3 online primer design software to design primer F11 (Forward primer: 5'-ATTCGCCCACCGATTCGTAT-3', Reversed primer 5'-GCCGGCGGAGAAGTTGTATA-3'), 20μl reaction system (ddH 2 O: 4.2μl; 2×Phanta Max buffer: 10μl; dNTPMix (10mM each): 0.4μl; Primer F (10μM): 1μl; Primer R (10μM): 1μl; Phanta maxsuper-fidelity DNAPolymerase (1U / μl): 0.4 μl; DNA (100ng / μl): 3μl), using PCR reaction program (98℃30s, {98℃15s, 62℃15s, 72℃3min}×7cycles, {98℃15s, 55℃15s, 72℃3min} ×22cycles, 72°C for 10min, stored at 16°C), amplified the BoFLC2 gene promoter region, and imaged by 1% agarose gel electrophoresis. The results are shown in figure 1 , the size of the amplified fragment of the promoter region of cabbage with early flowering and late flowering is about 2.0kb, and there is no difference in size;

[0059] (2) According to the BoFLC2 reference genome sequence, u...

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Abstract

The invention belongs to the field of molecular biology, and particularly relates to a molecular marker, a primer pair and a molecular marking method used for identifying the flowering time of brassica oleracea and application of the molecular marker, the primer pair and the molecular marking method. The molecular marker and the primer pair can be used for quick identification of a brassica oleracea breeding material and variety which are resistant to bolting and flower late and for the marker assisted selection of transfer breeding of the material which is resistant to bolting and flowers late, and are of great significance in brassica-oleracea-like vegetable breeding practice and theoretical research on bolting resistance.

Description

technical field [0001] The invention belongs to the field of molecular biology, and in particular relates to a molecular marker, a primer pair and a molecular marker method and application for identifying the flowering morning and evening of cabbage. Background technique [0002] Head cabbage (Brassica oleracea var.capitata), hereinafter referred to as cabbage, is a biennial vegetable crop of the cruciferous Brassica genus. In the process of vernalization in winter, in the spring and summer of the second year, flowering and fruiting are completed, and reproductive growth is completed. The edible organs of cabbage are mainly leaf bulbs, but in the production of cabbage, due to the poor bolting resistance of cabbage varieties and the frequent occurrence of cold in spring, cabbage will experience low temperature vernalization before the leaf bulbs mature. The phenomenon of immature bolting, once bolting occurs, will prevent the growth of leaf bulbs and directly bolt to bloom, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 宋洪元李勤菲李樟萍任雪松司军
Owner SOUTHWEST UNIVERSITY
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