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Primer set for hybrid cymbidium tortisepalum EST (expressed sequence tags)-SSR (simple sequence repeats) labeling and screening method

A screening method, the technology of Lily petal orchid, is applied in the primer set and screening field of EST-SSR marker of hybrid Lily petal orchid, it can solve the problems such as not seen, and achieve the effect of convenient statistics, simple operation of the method, and optimized primer ratio

Pending Publication Date: 2020-01-17
GUANGXI SUBTROPICAL CROPS RES INST GUANGXI SUBTROPICAL AGRI PROD PROCESSING RES INST
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  • Claims
  • Application Information

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Problems solved by technology

In recent years, with the expansion of the EST program in different species and the deepening of the research content, a large number of ESTs have been accumulated in many economically important plants and model plants, and the rapidly growing EST data provides a basis for the development of SSR markers. Rich sources, EST-SSR markers have been established in barley, rye, sugarcane, wheat, rice, kiwi, grape, cotton, spruce, apricot, alfalfa and other plants, but there is no research based on lotus The EST-SSR marker of the hybrid lotus petal orchid variety obtained by crossing the hybrid lotus petal orchid with other orchid varieties, so using the genetic data information of the hybrid lotus petal orchid plant variety to develop EST-SSR primers will have a greater impact on the genetics of the hybrid lotus petal orchid plant variety attitude and other research play a role in promoting

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  • Primer set for hybrid cymbidium tortisepalum EST (expressed sequence tags)-SSR (simple sequence repeats) labeling and screening method
  • Primer set for hybrid cymbidium tortisepalum EST (expressed sequence tags)-SSR (simple sequence repeats) labeling and screening method
  • Primer set for hybrid cymbidium tortisepalum EST (expressed sequence tags)-SSR (simple sequence repeats) labeling and screening method

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Experimental program
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Effect test

Embodiment 1

[0045] The screening method for the primer group marked by hybridization lotus petal orchid EST-SSR, it comprises the steps:

[0046] (1) Perform transcriptome sequencing on the RNA of the target hybrid orchid sample according to the conventional method to obtain the EST tag sequence, and then use MISA software to obtain the sequence segment where all the repeat sequences are located, and establish a repeat sequence database as the original sample of the target hybrid orchid orchid Gene database; the target hybrid lotus petal orchid sample is the blade of an annual plant;

[0047] (2) SSR site search is performed on the original gene database described in step (1), wherein the repeat numbers of one, two, three, four, five, and hexanucleotides are at least 10, 6, 5, 5, and 5 respectively ,5 times;

[0048] (3) Use the upstream and downstream sequences of the SSR site searched in step (2) as the target sequence, use Primer5.0 to design PCR primers for the target sequence, and g...

Embodiment 2

[0058] Embodiment 2: polymorphism analysis of hybrid lotus petal orchid EST-SSR marker

[0059] The polymorphism analysis was performed on the 4 pairs of primer pairs screened by the method described in Example 1 and used for hybridization of the EST-SSR marker of Lianpetalum orchid, and the results are shown in Table 1.

[0060] Table 1 Analysis table of polymorphism of EST-SSR markers in hybrid lotus petal orchid

[0061] Primer pair name Length (bp) total stripe polymorphic band polymorphism information content Primer pair A 248 73 6 0.563 Primer pair B 222 73 3 0.446 Primer pair C 174 75 3 0.4 Primer pair D 274 74 5 0.605

[0062] According to the above structural analysis, 4 pairs of primers can amplify 17 bands, and the number of polymorphic bands is between 3 and 6. On average, each pair of primers amplifies 4.25 polymorphic bands. The morphological ratios are all 100.00%. The polymorphism information content r...

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Abstract

The invention discloses a primer set for hybrid cymbidium tortisepalum EST (expressed sequence tags)-SSR (simple sequence repeats) labeling. The primer set includes 4 pairs of primers, and the nucleotide sequences of the primers are as shown in the sequence tables SED ID NO.1-SED ID NO.8. The invention further discloses a screening method of the primer set for hybrid cymbidium tortisepalum EST-SSRlabeling. The primer set for hybrid cymbidium tortisepalum EST-SSR labeling obtained by the screening method has the advantages of rich polymorphisms, stable amplification and convenient statistics,and can be used for genetic diversity analysis of germplasm resources of hybrid cymbidium tortisepalum plant varieties, molecular marker-assisted breeding and related molecular research.

Description

technical field [0001] The invention belongs to the technical field of molecular markers, and in particular relates to a primer set and a screening method for hybridizing EST-SSR markers of the lotus petal orchid. Background technique [0002] SSR (Simple Sequence Repeats) marking is a molecular marking technology based on specific primer PCR developed in recent years. 6) is a tandem repeat sequence of dozens of nucleotides consisting of repeating units, and the sequences on both sides of each SSR are generally relatively conserved single-copy sequences. Compared with other molecular markers, SSR markers have the following advantages: (1) rich in quantity, covering the entire genome, and revealing high polymorphisms; (2) having the characteristics of multi-allelic genes, providing high information; (3) Inherited in a Mendelian manner, showing co-dominance; (4) Each site is determined by the sequence of the designed primers, which is convenient for different laboratories to ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6858C12N15/11
CPCC12Q1/6895C12Q1/6858C12Q2600/156C12Q2531/113C12Q2565/125Y02A50/30
Inventor 罗清薄文浩李美育卢业飞覃茜丁丽琼陆祖正谢振兴秦玉燕周迎屈婷婷
Owner GUANGXI SUBTROPICAL CROPS RES INST GUANGXI SUBTROPICAL AGRI PROD PROCESSING RES INST
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