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Method for marking non-heading cabbage molecule by utilizing SAMPL technique

A technology of head cabbage and molecular markers, applied in the direction of biochemical equipment and methods, microbial determination/inspection, etc.

Inactive Publication Date: 2008-09-10
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

SAMPL has been applied to conifers, neem, Indian ginseng, lettuce, common wheat, cabbage, etc., but not yet applied to non-heading cabbage

Method used

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  • Method for marking non-heading cabbage molecule by utilizing SAMPL technique
  • Method for marking non-heading cabbage molecule by utilizing SAMPL technique
  • Method for marking non-heading cabbage molecule by utilizing SAMPL technique

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Cultivation of materials: Take 20 parts of non-heading Chinese cabbage from the Germplasm Resources Shanghai Branch of the State Key Laboratory as test materials (Table 1), select plump seeds, and sow them on a mixture of peat, vermiculite and organic fertilizer (volume ratio 4: 1). 4:1) in the mixed substrate, pour water thoroughly and drill a 0.5 cm deep hole, sow the seeds into the hole, and then cover with 0.5 cm thick peat. Seal it tightly with a black film and place it on a seedbed at about 30°C for 3-4 days. After germination and unearthed, transfer it to an artificial climate room with a day temperature of 25°C and a night temperature of 18°C ​​for 2 weeks. Leaves were used for DNA extraction.

[0032] DNA extraction: The CTAB method is used to extract DNA, and the specific operation steps are as follows:

[0033] 1. Leaf cleaning: select 3-5 young leaves, about 2g (grams), and wash them with pure water;

[0034] 2. Grinding leaves: freeze, add liquid nitrogen...

Embodiment 2

[0076] Cultivation of materials: 8 parts of non-heading Chinese cabbage with different disease resistance and strength from the Shanghai branch of the State Key Laboratory of Germplasm Resources were used as test materials (Table 2). Fertilizer (volume ratio 4: 4: 1) mixed substrate, after watering thoroughly, drill 0.5cm deep holes, sow seeds into the holes, and then cover with 0.5cm thick peat. Seal it tightly with a black film and place it on a seedbed at about 30°C for 3-4 days. After the seeds germinate and unearth, transfer them to an artificial climate room with a day temperature of 25°C and a night temperature of 18°C ​​for 2 weeks, and cut true leaves at the two-leaf stage. Leaves were used for DNA extraction.

[0077] DNA extraction: The CTAB method is used to extract DNA, and the specific operation steps are as follows:

[0078] 1. Leaf cleaning: select 3-5 young leaves, about 2g, and wash them with pure water;

[0079] 2. Grinding leaves: freeze, add liquid nitro...

Embodiment 3

[0117] Cultivation of materials: Take 5 parts of non-heading Chinese cabbage from the Germplasm Resources Shanghai Branch of the State Key Laboratory as test materials (Table 3), select plump seeds, and sow them on a mixture of peat, vermiculite and organic fertilizer (volume ratio 4: 1). 4:1) in the mixed matrix, after pouring water thoroughly, drill a 0.5cm deep hole, sow the seeds into the hole, and then cover with 0.5cm thick peat. Seal it tightly with a black film and place it on a seedbed at about 30°C for 3-4 days. After the seeds germinate and unearth, transfer them to an artificial climate room with a day temperature of 25°C and a night temperature of 18°C ​​for 2 weeks, and cut true leaves at the two-leaf stage. Leaves were used for DNA extraction.

[0118] DNA extraction: The CTAB method is used to extract DNA, and the specific operation steps are as follows:

[0119] 1. Leaf cleaning: select 3-5 young leaves, about 2g, and wash them with pure water;

[0120] 2. G...

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Abstract

The invention relates to a method for loose head cabbage molecular markers by utilizing the SAMPL technology comprising DNA extraction, DNA enzyme digestion-connection system optimization, SAMPL pre-amplification system optimization, SAMPL amplification system optimization, electrophoresis, silver staining color development and cluster analysis, wherein, a DNA sample is performed enzyme digestion through using EcoRI, MseI or PstI enzyme, and then corresponding joints of EcoRI,MseI or PstI are connected on the DNA sample through using T4-DAN ligase; the SAMPL pre-amplification is performed through using a 10mu L reaction system; a pre-amplification primer is corresponding to the chosen enzyme in the enzyme digestion; the SAMPL amplification is performed through using a 20mu L reaction system, one amplification primer is corresponding to the pre-amplification primer, and the other is an SAMPL primer. The method for loose head cabbage molecular markers by utilizing the SAMPL technology performs optimization and primer design for the complicated system of the SAMPL, constructs an SAMPL reaction system applicable to loose head cabbages and realizes loose head cabbage molecular markers.

Description

technical field [0001] The invention relates to a molecular marker method in the technical field of crop breeding, in particular to a method for molecular marker of non-heading Chinese cabbage by using SAMPL (selectively amplified polymorphic microsatellite site) technology. Background technique [0002] Second-generation molecular markers based on PCR (polymerase chain reaction) technology, such as AFLP (amplified fragment length polymorphism) and SSR (simple sequence repeat), have become important technical means of molecular marker-assisted selection. As the combination of modern molecular biology and traditional genetic breeding, molecular marker-assisted selection technology can select breeding materials at the DNA level with the help of molecular markers, speed up the selection process, and enhance the accuracy of selection, thereby significantly improving breeding efficiency. So the second generation molecular marker technology has been widely used in breeding work. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 陈火英王新华刘杨
Owner SHANGHAI JIAO TONG UNIV
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