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Multi-combination molecular markers related to resistance of litopenaeus vannamei and application

A resistance-related, molecular marker technology, applied in recombinant DNA technology, microbial determination/inspection, biochemical equipment and methods, etc., to achieve the effect of improving efficiency and accuracy, and being less susceptible to environmental influences

Active Publication Date: 2016-10-26
INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most of the previous reports were about the application of a single marker in resistance breeding, and the combined assisted selection of multiple markers has not been reported yet.

Method used

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  • Multi-combination molecular markers related to resistance of litopenaeus vannamei and application
  • Multi-combination molecular markers related to resistance of litopenaeus vannamei and application
  • Multi-combination molecular markers related to resistance of litopenaeus vannamei and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Obtainment of WSSV resistance marker combination of Litopenaeus vannamei

[0037] 1. Acquisition of WSSV susceptible and resistant materials

[0038] WSSV susceptible and resistant materials come from the mixed population of "Kehai No. 1" multi-lineage. 500 healthy Litopenaeus vannamei were selected and placed in 12 50L aquariums. Before the experiment, 10 prawns were randomly selected to detect no WSSV infection. The experimental water temperature was (23±1)℃. After the starvation treatment, the diseased shrimps caused by WSSV infection were fed (WSSV virus content was 10 4 -10 6 copy / ng DNA). After the challenge, check 3 times a day (each interval 6h) and collect dead shrimp, randomly sample and extract genomic DNA, and the earliest dead 48 prawns are used as the WSSV susceptible group. On the 14th day after challenge, the survival of Litopenaeus vannamei was stable, and 48 surviving prawns were selected as the WSSV-resistant group.

[0039] 2. Genomic...

Embodiment 2

[0080] Example 2: Application of Litopenaeus vannamei WSSV Resistance Marker Combination

[0081] Based on the results of the above association analysis, this study established a method of using the obtained molecular marker combination to assist in the selection of disease resistance. The specific operations are as follows:

[0082] 1. According to the instructions of the TIANGEN Plant Genomic DNA Extraction Kit, extract the individual DNA of the breeding material, use the NanoDrop 1000 to detect the DNA concentration and purity of the extracted DNA, and use the tested DNA concentration > 50ng / μl and purity OD 260 / 280 The individual DNA of 1.7 to 1.9 was detected by 1% agarose gel electrophoresis, and the individuals with complete DNA fragments as a result of electrophoresis were selected for use.

[0083] 2. SNP typing of three markers:

[0084] Individuals obtained in the above-mentioned manner are subjected to SNP typing by using the above-mentioned embodiment to obtain t...

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Abstract

The invention belongs to the technical field of germplasm innovation of aquatic animals and particularly relates to a method for for breeding litopenaeus vannamei with WSSV (white spot syndrome virus) resistance on the basis of multi-marker combination assistance and an application of the multi-combination markers. SNP marker typing is performed on SNP loci of immune signaling pathways of the litopenaeus vannamei in sensitive groups and resistance groups with a high throughput typing technology, and 6 SNP molecular markers in significant correlation with WSSV resistance are obtained. The optimal resistance maker combination is obtained through screening with a multi-marker interaction analysis method and comprises nLvALF2g-89-A>G, Unigene34569g-97-T>C and TRAF6g-139-A>G, and four genotype combinations including AA-TT-AA, AA-CT-AA, AA-CT-AG and AG-CT-AA of the three markers show an obvious correlation advantage with the WSSV resistance. Candidate markers are provided for disease-resistant molecular breeding of prawns, one novel path is provided for molecular marker assistant resistance breeding of the prawns, and broad application prospect is realized in research of WSSV-resistance variety breeding of the prawns.

Description

technical field [0001] The invention belongs to the technical field of aquatic animal germplasm innovation, and in particular relates to a multi-combination molecular marker related to the resistance of Litopenaeus vannamei. Background technique [0002] Litopenaeus vannamei is the prawn species with the largest farming scale and output in the world. Since it was introduced into China in 1988, the output of Litopenaeus vannamei has increased rapidly, and it has become the most important aquaculture species in my country. However, with the expansion of farming scale, disease outbreaks are becoming more and more frequent. Among them, WSSV has become the most serious disease affecting the shrimp industry in the world due to its high lethality and rapid spread. [0003] Breeding new disease-resistant varieties is an important means to solve the problem of shrimp diseases. Compared with traditional breeding methods, modern breeding technology combined with molecular marker-assis...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/156
Inventor 李富花于洋刘敬文张晓军相建海
Owner INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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