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Microsatellite Primer Combination and Application for Identification of Genetic Relationship in Eggs of Green-tailed Pheasant's Pheasant

A green-tailed iridescent pheasant and microsatellite marker technology, which is applied in the field of DNA molecular markers in molecular biology, can solve problems such as degradation and less DNA, and achieve the effects of increasing accuracy, facilitating management, and reducing the risk of inbreeding

Active Publication Date: 2022-06-07
CHENGDU RES BASE OF GIANT PANDA BREEDING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The DNA extracted from this non-destructive sampling sample has the problem of small amount of DNA and degradation, which also puts forward higher requirements for the screening of microsatellite markers

Method used

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  • Microsatellite Primer Combination and Application for Identification of Genetic Relationship in Eggs of Green-tailed Pheasant's Pheasant
  • Microsatellite Primer Combination and Application for Identification of Genetic Relationship in Eggs of Green-tailed Pheasant's Pheasant
  • Microsatellite Primer Combination and Application for Identification of Genetic Relationship in Eggs of Green-tailed Pheasant's Pheasant

Examples

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Effect test

Embodiment 1

[0040] The present invention in the whole genome sequence of green-tailed rainbow pheasant, microsatellite site development, while designing primers of these sites, selecting several nucleotide sequences containing microsatellite repeat sites, species-specific screening, selecting 200 pairs of species-specific primer sequences to synthesize common primers, PCR of randomly selected 3 green-tailed rainbow pheasant DNA samples, and electrophoresis on agarose gel, primer initial screening, to obtain 93 microsatellite sites, on the basis of which 93 pairs of fluorescent primers were synthesized, PCR amplification and genotyping were performed in multiple DNA samples, and the screening of green-tailed pheasant microsatellite sites was completed, and 12 pairs of molecular markers with high polymorphism and good reproducibility were obtained. The specific content is as follows:

[0041] 1) Extraction of green-tailed rainbow pheasant DNA

[0042] During the breeding period of the green-tai...

Embodiment 2

[0057] QiAMP 69506 Blood / Tissue Trace Kit Operation Manual was used to extract a total of 6 parts of eggshell (hatching completed or stillborn) DNA in three enclosures in the captive population of Green-tailed Rainbow Pheasant in 2018, and the extraction method was the same as Example 1. 12 pairs of microsatellite primers were used (Table 1). PCR reaction conditions are the same as In Example 1, each reaction is repeated more than 3 times to ensure consistency of results.

[0058] Table 1 Primers

[0059]

[0060]

[0061] The above PCR products were genotyped using a generation gene sequencer, and the microsatellite typing data (amplification fragment length) are shown in Table 2.

[0062] Table 2 Microsatellite typing data (amplified fragment length bp)

[0063]

[0064] The above 6 samples were identified by software ML-Relate, and the analysis results are shown in Table 3. HS (Half Sibling) represents a half-cell relationship, FS (Full Sibluing) represents a whole cell r...

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Abstract

The invention discloses a combination and application of microsatellite primers for identification of phylogenetic relationship of birds' eggs in the same house. The primers include CM34, CM104, CM412, CM687, CM1423, CM1959, CM2014, CM2083, CM2269, CM2289 and CM3131 and CM3590, the nucleotide sequences of which are shown in SEQ ID NO.1-24. The primers of the present invention can be stably amplified in the eggshell DNA of the green-tailed rainbow pheasant. Through this group of 12 pairs of primers, the microsatellite sequence of the green-tailed rainbow pheasant with high polymorphism content and good repeatability is obtained, and the corresponding markers of these markers are established. Genotyping technology, and can realize the identification of siblings, half-parents or non-parents among laying eggs in the same house of green-tailed rainbow pheasant.

Description

Technical field [0001] The present invention belongs to the field of molecular biology DNA molecular labeling technology, specifically relates to a green-tailed rainbow pheasant and bird egg affinity identification of microsatellite primer combinations and applications. Background [0002] The green-tailed rainbow pheasant is a rare bird endemic to China, and like the giant panda, it was found and named in Baoxing County. Distributed in the inaccessible alpine areas of Sichuan, Yunnan, Qinghai, Gansu and Tibet, the green-tailed rainbow pheasant has high ecological protection value and is also an umbrella species in alpine meadow ecosystems. The green-tailed pheasant is rare in nature, and in 2016, a team of experts at IUCN estimated its adult population to be between 2,500 and 9,999, and speculated that wild populations are still in the process of continuous decline. At present, the green-tailed rainbow pheasant is listed in Appendix I of the Washington Convention (effective date...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12Q1/6858C12N15/11
CPCC12Q1/6888C12Q1/6858C12Q2600/156C12Q2531/113C12Q2565/125C12Q2563/107Y02A50/30
Inventor 张亮侯蓉岳碧松沈富军吴里霞陈黎安俊辉张修月张文平寇洁王也李严刘佳文王涓刘红
Owner CHENGDU RES BASE OF GIANT PANDA BREEDING
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