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CMS molecular tag based on hibiscus cannabinus mitochondrial gene non-coding region and primer pair and application of CMS molecular tag

A mitochondrial gene, non-coding region technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve problems such as single cytoplasmic type of kenaf and large-scale occurrence of diseased physiological races

Active Publication Date: 2021-06-01
GUANGXI ZHUANG AUTONOMOUS REGION ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the single type of kenaf cytoplasm, it is easy to cause the potential risk of a large-scale occurrence of a certain disease physiological race. On the other hand, the advantages and resistance of hybrids have obvious cytoplasm effects. Therefore, a variety of sterile cytoplasms have been created. Germplasm is the key problem to realize the sustainable utilization of kenaf heterosis

Method used

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  • CMS molecular tag based on hibiscus cannabinus mitochondrial gene non-coding region and primer pair and application of CMS molecular tag
  • CMS molecular tag based on hibiscus cannabinus mitochondrial gene non-coding region and primer pair and application of CMS molecular tag
  • CMS molecular tag based on hibiscus cannabinus mitochondrial gene non-coding region and primer pair and application of CMS molecular tag

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] Extraction of Total DNA from Kenaf

[0079] The tips, mortar, and extraction buffer used in the extraction process of kenaf total genomic DNA were first autoclaved for 20 minutes, and then taken out for later use. Extract DNA according to the following steps:

[0080] (1) Preheat CTAB (0.1 mol / L Tris-HCl pH8.0, 0.025 mol / L EDTA-Na2pH8.0, 1.4 mol / LNaCl, 2% CTAB (w / v)) extract in a 65°C water bath;

[0081] (2) Grind 2 to 3 g of fresh or -80°C frozen leaves in liquid nitrogen;

[0082] (3) Quickly add CTAB extract, mix well, pour into a centrifuge tube, and place in a water bath at 65°C for 30 min. gently shaking in between;

[0083] (4) Add an equal volume of chloroform / isoamyl alcohol, mix well, and centrifuge at 12000 rpm for 10 min at room temperature;

[0084] (5) Take the supernatant, add 5M NaCl and absolute ethanol according to the ratio of extract: 5M NaCl: dehydrated ethanol = 1:1:1 to prepare a mixture, invert and mix well;

[0085] (5) Aspirate the superna...

Embodiment 2

[0099] Application of HMBQ102 Marker in Breeding Kenaf Cytoplasmic Male Sterile Line

[0100] Using this molecular marker to detect the existing known kenaf cytoplasmic male sterile line and maintainer line materials in our research group, the results are as follows figure 2 The size of the band amplified by the molecular tag in the known cytoplasmic male sterile lines of kenaf with different cytoplasmic types is 980bp (lane 1-10), and the band size amplified in the maintenance is 1082bp (lane 12- 19), indicating that this molecular signature can accurately identify the cytoplasmic male sterile line resources of different cytoplasmic types of kenaf.

Embodiment 3

[0102] Repeat experiment for stability of HMBQ102 labeling

[0103] According to the stable characteristic that the kenaf HMBQ102 molecular tag can amplify a band with a size of 980bp in the sterile cytoplasm and a band with a size of 1082bp in the maintainer line, the field planting resources of kenaf were screened and identified. The result is as image 3 Shown: In this example, a total of 16 germplasm resources with consistent cytoplasmic characteristics with UG93A and 10 germplasm resources with consistent cytoplasmic characteristics with UG93B were identified. The identification of the above materials provides a molecular basis for the selection and breeding of kenaf cytoplasmic androgenic lines. On the other hand, UG93BS1 and UG93BS2 ( image 3 a Lane 5, Lane 6) These two germplasm resources are two sterile individuals found in the maintainer line UG93B population. HMBQ102 molecular label detection shows that these two individuals have the same cytoplasmic type as the ...

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Abstract

The invention belongs to the technical field of biology, and relates to hibiscus cannabinus breeding, in particular to a CMS molecular tag based on a hibiscus cannabinus mitochondrial gene non-coding region and a primer pair and application of the CMS molecular tag. The CMS molecular tag is named as HMBQ102 based on deletion of 1 bp and 101 bp at 535 bp and 995 bp of an upstream non-coding region of a mitochondrial gene cob respectively. The CMS molecular tag based on a kenaf mitochondrial gene non-coding region and the primer pair and application of the CMS molecular tag are developed on the basis of the difference between the kenaf cytoplasmic male sterile line mitochondrial genome and the kenaf cytoplasmic male sterile line mitochondrial genome, the tag is easy to operate, kenaf male sterile cytoplasm can be rapidly and accurately identified, and the tag has an important application value in the breeding and heterosis utilization of the kenaf cytoplasmic male sterile line.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to kenaf breeding, in particular to a CMS molecular tag based on the non-coding region of the kenaf mitochondrial gene and its primer pair and application. Background technique [0002] Cytoplasmic male sterility (CMS) is one of the main means of kenaf hybrid breeding. However, due to the single type of kenaf cytoplasm, it is easy to cause the potential risk of a large-scale occurrence of a certain disease physiological race. On the other hand, the advantages and resistance of hybrids have obvious cytoplasm effects. Therefore, a variety of sterile cytoplasms have been created. Germplasm is the key problem to realize the sustainable utilization of kenaf heterosis. Molecular marker-assisted breeding is an important means to speed up the selection of cytoplasmic male sterile lines of kenaf. [0003] The innovative team of Professor Zhou Ruiyang of Guangxi University discovered a male steril...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 廖小芳侯文焕唐兴富赵艳红
Owner GUANGXI ZHUANG AUTONOMOUS REGION ACAD OF AGRI SCI