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A wheat stalk breaking strength molecular marker qwqd4b.4-13 and its application

A breaking strength and molecular marker technology, applied in the field of genetic engineering, can solve the problems of false positive QTLs, QTLs of stem breaking strength that have not been verified by the breeding process or variety validity, and poor accuracy.

Active Publication Date: 2019-10-29
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, few genes related to wheat stalk breaking strength have been identified, and there is a lack of effective markers for molecular marker-assisted selection. The main reason is that it is limited by the following two aspects. Verified by the breeding process or variety effectiveness; on the other hand, due to the loci mapped by a single genetic population, the accuracy is poor, and due to the large confidence interval, too many QTLs or the existence of false positive QTLs, the stem break strength is reduced Molecularly assisted The actual efficiency of selection cannot even be effectively applied to wheat breeding

Method used

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  • A wheat stalk breaking strength molecular marker qwqd4b.4-13 and its application
  • A wheat stalk breaking strength molecular marker qwqd4b.4-13 and its application
  • A wheat stalk breaking strength molecular marker qwqd4b.4-13 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] The leaf DNA of embodiment 1 wheat is extracted

[0020] (1) Take 0.3-0.5g leaves into a 5mL centrifuge tube, freeze them in liquid nitrogen and grind them into powder;

[0021] (2) Add 1600 μL of buffer S that has been preheated to 65°C, mix by inversion several times, bathe in water for 0.5-1 hour, and shake gently several times during this period to fully mix;

[0022] (3) Cool down to room temperature, wait for 10 minutes, add 10-15μL RNase (10mg / mL) in a 37℃ water bath for 30 minutes, and shake gently several times to fully mix, about once / 10min;

[0023] (4) Take out the centrifuge tube, add an equal volume of 1600 μL, 4°C phenol (Tris-balanced phenol): chloroform: isoamyl alcohol (volume ratio 25:24:1) for extraction, mix gently for 10 minutes, and stand in the refrigerator at 4°C 5min, then centrifuge at 8000rpm for 10min;

[0024] (5) Take the supernatant in another tube, about 1300 μL, add an equal volume of cold chloroform (placed in a refrigerator at 4°C),...

Embodiment 2

[0040] Embodiment 2 target product amplification

[0041] Forward primer sequence: ggtggttcct ttcgattttg cgc (as shown in SEQ ID NO:4)

[0042] Reverse primer sequence: tgttacggta cacaaactca ctagt (as shown in SEQ ID NO:5)

[0043] PCR amplification: the PCR amplification system is 20μL

[0044]

[0045] Note: Mix available: or (Taq enzyme 0.25μL, DNK 2.0μL, Buffer1.5μL, MgCl0.4μL configuration)

[0046] Amplification conditions:

[0047]

[0048] A 721bp fragment can be obtained through the above amplification, and its nucleotide sequence is shown in SEQ ID NO:1.

Embodiment 3

[0049] Example 3 Specific enzyme digestion of PCR products:

[0050] Enzyme digestion system 10μL:

[0051] Specific enzyme AluI: 0.3 μL

[0052] PCR product: 2 μL

[0053] 10×NE buffer: 1.0μL

[0054] wxya 2 O: 6μL

[0055] Enzyme digestion reaction conditions: add 0.3 μL AluI specific enzyme (commercially available) to the PCR amplification product, bathe in water at 37°C for 4 hours, then inactivate the enzyme digestion system at 65°C for 5 minutes.

[0056] After electrophoresis and separation of the above-mentioned amplified products on 8% polyacrylamide gel, the molecular weight of the amplified product is 721bp, and then the corresponding fragment is detected after digestion with AluI-specific enzyme. A fragment of 721bp proves that the fragment has not been cut, and the variety (line) contains the gene locus QWQD4B.4-13 for enhancing the stem breaking strength. If it contains nucleotide sequences such as SEQ ID NO:2 and 3 after detection The above two fragments p...

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Abstract

A wheat stalk breaking strength molecular marker is disclosed, the wheat stalk breaking strength molecular marker is located between wheat 4B chromosome TDURUM-CONTIG63670-287 and IACX557, and named as QWQD4B.4-13; through application of the molecular marker, whether QWQD4B.4-13 gene loci for increasing stalk breaking strength exist in wheat varieties or lines can be detected, the breeding process of lodging-resistant wheat varieties can be accelerated, by use of the special stalk breaking strength molecular marker, the screening is fast and accurate, and is not affected by the environment, a target is clear to select, cost of production is saved, the selection efficiency and quality of the lodging-resistant high yield wheat varieties or lines are greatly improved, QTL detection effect and precision can be improved, the effective molecular marker is developed, a stalk breaking strength molecular assisted breeding system is developed, and the stalk breaking strength molecular assisted breeding system can be programmatically applied in breeding practices.

Description

Technical field: [0001] The invention relates to the technical field of genetic engineering, can be applied to the field of wheat yield breeding, and specifically provides a wheat stalk breaking strength molecular marker and its application. Background technique: [0002] Wheat is the second largest grain crop in my country, and more than 1 / 3 of the world's population uses wheat as a staple food. Lodging is a common problem in wheat production. Wheat after lodging not only reduces yield and is inconvenient to harvest, but also seriously affects product quality, resulting in thin grains, reduced bulk density, poor milling quality, and reduced wheat commerciality and manufacturability. , directly affects the production and processing of high-quality wheat. Wheat lodging generally occurs at 10%-30% of the base of the stalk, that is, the second and third nodes of the stalk base. The breaking strength of the wheat stalk is a comprehensive indicator of the lodging resistance of w...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/124
Inventor 田纪春陈建省刘凯谢楚鹏王芳芳孙晓晓张海军安玉玲刘佟佟邵文张若晴
Owner SHANDONG AGRICULTURAL UNIVERSITY