Intron 3-based Molecular Marker for Identifying Barley Semi-dwarf Multi-tiller Gene fol-a and Its Application

A fol-a, molecular marker technology, applied in the field of agricultural biology, can solve problems such as environmental pollution, increased fertilizer input, and increased production costs

Active Publication Date: 2021-09-14
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Semi-dwarf crops increase the lodging resistance of crops, but at the same time increase the input of fertilizers, leading to problems such as increased production costs and environmental pollution

Method used

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  • Intron 3-based Molecular Marker for Identifying Barley Semi-dwarf Multi-tiller Gene fol-a and Its Application
  • Intron 3-based Molecular Marker for Identifying Barley Semi-dwarf Multi-tiller Gene fol-a and Its Application
  • Intron 3-based Molecular Marker for Identifying Barley Semi-dwarf Multi-tiller Gene fol-a and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1, design molecular marker SNPE3

[0042]1), test materials: mutants BW370 and Bowman;

[0043] The gene fol-a controlling semi-dwarf and multi-tiller traits was cloned from the mutant BW370, and the nucleotide sequence of the gene is as described in SEQ ID NO:1. The protein encoded by the semi-dwarf multi-tiller gene fol-a has the amino acid sequence described in SEQ ID NO:3.

[0044] The radiation-induced mutant Proctor (the mutant has semi-dwarf and multi-tiller traits) was backcrossed with barley variety Bowman multiple times to obtain mutant BW370, which has Bowman's genetic background but exhibits semi-dwarf and multi-tiller traits.

[0045] The nucleotide sequence of the wild-type Bowman HORVU2Hr1G098820 gene is described in SEQ ID NO: 2, and the protein encoded by it has the amino acid sequence described in SEQ ID NO: 4.

[0046] 2) Obtaining the molecular marker of the semi-dwarf and multi-tiller gene fol-a: According to the sequence comparison res...

Embodiment 2

[0052] Embodiment 2, identify the sequence difference of mutant BW370 and Bowman with molecular marker:

[0053] 1), DNA extraction: Take 100 mg of barley leaves, and use the CTAB method to extract DNA. The specific steps are as follows:

[0054] ① Grind the sheared leaves in liquid nitrogen and put them into 1.5ml centrifuge tubes, add 600 μL of CTAB to each tube and place in a warm water bath at 65°C for 50-60 minutes (take out every 10 minutes and shake gently);

[0055] ②Add 600 microliters of chloroform / isoamyl alcohol (24:1) and mix well for 15 minutes. Slowly, weigh in pairs, and after balancing, centrifuge at 9600 rpm for 10 minutes at 4°C;

[0056] ③Pipe the supernatant to another 1.5ml new centrifuge tube, add 2 times the volume of absolute ethanol and mix slightly (precipitate in -20°C refrigerator for 30-60min);

[0057] ③ Pick out the precipitate with a pipette tip, put it in a 1.5ml centrifuge tube, wash it twice with 70% ethanol and air dry;

[0058] ④ Add 100...

Embodiment 3

[0067] Example 3. Using the molecular marker SNPE3 to detect 32 barley germplasm resources (including mutants BW370 and Bowman, 2 repetitions) according to the method described in Example 2, the barley germplasm resources include wild barley, local varieties and from all over the world The barley cultivars and the F 1 (See Table 1);

[0068] The result is as Figure 4 shown.

[0069] The test results show that the F of BW370 / Vlamingh in these materials 1 The hybrid is heterozygous, and the mutant BW370 is the mutant genotype; other materials are all wild-type genotypes, and their phenotypes are also different from the semi-dwarf multi-tiller phenotype of the mutant BW370, indicating that they are not Containing the Hvthd mutant gene also proves that this molecular marker can be used in the detection of barley germplasm resources.

[0070] Table 1. Information on 32 barley germplasm resources detected by molecular marker SNPE3

[0071]

[0072]

[0073] Verification ...

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Abstract

The invention discloses a gene fol-a controlling semi-dwarf and multi-tiller traits, the nucleotide sequence of the gene is as described in SEQ ID NO:1. The present invention also discloses the protein encoded by the above-mentioned semi-dwarf multi-tiller gene fol-a, the amino acid sequence of which is as described in SEQ ID NO:3. The invention also discloses a molecular marker SNPE3 based on intron 3 for identifying the barley semi-dwarf multi-tiller gene fol-a. Using the molecular marker SNPE3, it is possible to identify whether the fol‑a semi-dwarf multi-tiller gene is present in barley. Using the marker to perform semi-dwarf multi-tiller breeding assisted selection of the fol-a gene can guarantee 100% accuracy and speed up the progress of barley semi-dwarf multi-tiller breeding.

Description

technical field [0001] The invention relates to a gene marker method for identifying barley semi-dwarf multi-tiller gene fol-a and corresponding primers. The method belongs to the field of agricultural biotechnology and can be used for rapid screening of barley semi-dwarf multi-tiller germplasm and fol-a-containing Molecular marker-assisted breeding of mutant genes. Background technique [0002] Many factors such as population growth, climate change and natural environment damage have led to global food shortages, and how to increase crop yields has become a common concern of breeders. In the 1960s, the application of semi-dwarf genes significantly increased crop yields, which was called the "Green Revolution" (Peng et al., 1999; Monna et al., 2002), and the "Green Revolution Gene" involved Gibberella pathways of metabolism. Semi-dwarf crops increase the lodging resistance of crops, but at the same time, they also increase the amount of fertilizer input, resulting in incre...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12Q1/6895C12N15/11
CPCC07K14/415C12Q1/6895C12Q2600/13C12Q2600/156
Inventor 华为谈聪汪军妹朱靖环尚毅李承道杨建明巫小建
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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