Unlock instant, AI-driven research and patent intelligence for your innovation.

A kind of preparation method of detection sample of wheat seed high molecular weight glutenin subunit

A gluten subunit, high molecular weight technology, applied in the field of grain chemistry, to achieve the effects of low labor consumption, high efficiency and simple operation

Active Publication Date: 2017-06-23
淮安明天种业科技有限公司
View PDF4 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Related methods have not yet been reported

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A kind of preparation method of detection sample of wheat seed high molecular weight glutenin subunit
  • A kind of preparation method of detection sample of wheat seed high molecular weight glutenin subunit
  • A kind of preparation method of detection sample of wheat seed high molecular weight glutenin subunit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: Reagent preparation

[0030] (1) Preparation of related reagents for sample extraction

[0031] Soaking reagent A: 25% n-propanol (v / v) + 0.04M Tris-HCl (PH8.0)

[0032] Extraction reagent B: 25% n-propanol (v / v) + 0.04M Tris-HCl (PH8.0) + 3.0% SDS (w / v, sodium dodecyl sulfate) + 1.3% DTT (v / w, Dithiothreitol) + 2.0% 4-VP (v / v, 4-vinylpyridine). Among them, SDS, DTT and 4-VP are added in proportion and shaken well before sample extraction.

[0033] (2) Preparation of related reagents for polypropylene gel electrophoresis

[0034] Gel preparation related solutions: 1M Tris-HCl buffer (pH 8.8); 1M Tris-HCl buffer (pH6.8); Acr:Bis aqueous solution (394.8gAcr+5.2g Bis, dilute to 1000mL); 10% SDS (w / v) in water; 1.5% ammonium persulfate in water (w / v).

[0035] 5× electrode buffer: 1.5% Tris (w / v) + 7.2% g glycine (w / v) + 0.5% (w / v) SDS, distilled water to volume.

[0036] Fixative: 12% trichloroacetic acid (w / v) in water.

[0037] Staining solution: 40%...

Embodiment 3

[0039] Embodiment 3: the embryo end sample cutting of single seed

[0040] Select 23 production promotion varieties or control varieties, as shown in Table 2 below. Cut 1 / 4 to 1 / 5 of the seed volume from the embryoless end of each single seed, and put them into a 96-well microplate in turn, and put them in the matrix registration form consistent with the position of the microwells of the 96-well microplate Record the sample name, and then put a 2mm steel ball in each hole.

[0041] Table 2 HMW-GS identification results of 23 seed materials

[0042]

[0043]

Embodiment 4

[0044] Example 4: High Molecular Weight Glutenin Subunit (HMW-GS) Extraction

[0045] 1. Use an Eppendorf 100uL eight-channel pipette to add 40uL of reagent A to the 96-well microplate where the cut seeds and steel beads have been placed, cover with a 12-well plastic microwell cover, and let stand overnight.

[0046] 2. Open the stripped microwell cover, add 120uL of reagent B to the 96-well microplate with an eight-channel pipette, and cover the 12-well plastic microwell cover.

[0047] 3. Put the microplate into a high-throughput tissue mill (Retsch TissueLyser), the milling frequency is 30 times / s, and the milling time is 2min, and then put the microplate into a dry heat blast oven at 60°C Stand still, during every 14 minutes of standing, vortex once for 1 minute, twice for a total of 30 minutes.

[0048] 4. Put the incubated microwell plate into an Eppendorf 5810R desktop centrifuge and centrifuge at 4000rpm×15min.

[0049] 5. Use an eight-channel pipette to pipette 20uL...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
diameteraaaaaaaaaa
Login to View More

Abstract

The invention relates to a preparation method of a detection sample of a high-molecular-weight subunit of glutenin of a wheat seed and belongs to the biochemical field. The preparation method comprises the following steps: slitting 1 / 4-1 / 5 of the wheat seed by volume at an embryo free end; putting the wheat seed in a 96-hole microporous plate for PCR amplification with the single pore volume of 200 microliter; putting a steel ball with the diameter of 2mm into each micropore; then adding 40 microliter of a soak buffer liquid into the microporous plate for standing overnight; then adding 120 microliter of an extraction buffer liquid; grinding by a high flux grinder, then standing for 30 minutes in an oven at 60 DEG C, and carrying out vortex shaking for 1 minute twice in the period; and centrifugalizing the microporous plate to obtain a supernate which is the sample required for detecting the high-molecular-weight subunit of glutenin of wheat. The extraction method is high in flux and suitable for preparing samples for detecting the high-molecular-weight subunit of glutenin of the wheat seed on a large scale.

Description

technical field [0001] The invention relates to a preparation method of a wheat seed high molecular weight glutenin subunit detection sample, belongs to the field of grain chemistry, and is specially used for the chemical extraction of wheat glutenin. Background technique [0002] High molecular weight glutenin subunit (High molecular weight glutenin subunit, HMW-GS) plays a decisive role in the quality of wheat gluten (Gianibelli et al., 2001). Gene coding, but most varieties have 3 to 5 HMW-GS. Excavating and aggregating high-quality HMW-GS is the most important approach for wheat quality breeding at home and abroad, so how to quickly extract and identify HMW-GS will directly affect the effectiveness of molecular breeding for wheat gluten quality. [0003] Due to the additive and interactive effects of HMW-GS on gluten strength, multiple subunits must be tracked simultaneously in a breeding program involving 2-3 HMW-GS crosses. Polymerase chain reaction (PCR) of a single...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/28
Inventor 张平平马鸿翔姚金保张鹏杨丹王化敦
Owner 淮安明天种业科技有限公司