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SSR molecular marker primer capable of rapidly identifying robinia pseudoacacia polyploids

A technology of molecular markers and polyploids, applied in the determination/testing of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problem of rapid identification of Robinia pseudoacacia polyploids, which cannot be quickly identified High-efficiency SSR primers and other issues to achieve high polymorphism and good repeatability

Inactive Publication Date: 2018-03-06
SHANDONG FOREST SCI RES INST +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, at present, there is no method for rapid identification of Robinia pseudoacacia polyploids using fluorescent SSR labeling technology at home and abroad, and there is no efficient SSR primer that can quickly identify Robinia pseudoacacia polyploids

Method used

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  • SSR molecular marker primer capable of rapidly identifying robinia pseudoacacia polyploids
  • SSR molecular marker primer capable of rapidly identifying robinia pseudoacacia polyploids
  • SSR molecular marker primer capable of rapidly identifying robinia pseudoacacia polyploids

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Experimental program
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Effect test

Embodiment 1

[0028] The SSR molecular marker primers for rapid identification of Robinia pseudoacacia polyploids were obtained through the following steps:

[0029] (1) Extraction of Robinia pseudoacacia total RNA and transcriptome sequencing

[0030] Total RNA was extracted using Trizol reagent. The pipette tip, mortar and spoon used in the experiment were all treated with DEPC (diethyl pyrocarbonate) for 4 hours, followed by high temperature and high pressure sterilization for 1 hour. Then, total RNA was extracted according to the usage method of Trizol reagent. Use 1.0% non-denaturing agarose gel electrophoresis to roughly detect the integrity of the extracted RNA; use the instrument Nanodrop (IMPLEN, CA, USA) to detect the purity and concentration of the extracted RNA (OD 260 / 230 ratio; OD 260 / 280 ratio) ; RNA integrity was detected using Agilent 2100 (Agilent Technologies, CA, USA).

[0031] After the RNA sample is qualified, in order to ensure high-quality transcriptome data, it w...

Embodiment 2

[0048] 1. Test sample

[0049] In this experiment, five tetraploid Robinia pseudoacacia clones introduced from Korea were used as test materials. They are K1 (Tetraploid R. pseudoacacia K1), K2 (Tetraploid R. pseudoacacia K2), K3 (Tetraploid R. pseudoacacia K3), K4 (Tetraploid R. pseudoacacia K4), and K5 (Tetraploid R. pseudoacacia K5).

[0050] The materials were collected from Daqingshan Forest Farm, Fei County, Shandong Province.

[0051] In May 2016, the young leaves were collected, dried on silica gel, and set aside.

[0052] 2. Genomic DNA extraction:

[0053] Take the young leaves of Robinia pseudoacacia clones, and use the plant tissue genomic DNA extraction kit (magnetic bead method) (Cat. No. PTED-6030) of Inrich Biochemical Technology (Shanghai) Co., Ltd. to extract according to the instructions.

[0054] 3. Use SSR primers for PCR amplification:

[0055] Using the total DNA sample of Robinia pseudoacacia extracted in step 2 as a template, use the following two ...

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Abstract

The invention discloses an SSR molecular marker primer capable of rapidly identifying robinia pseudoacacia polyploids, wherein the primer base sequence is as shown in SEQ ID NO.1 and SEQ ID NO.2. According to the invention, robinia pseudoacacia polyploids can be rapidly identified from the molecular level without being affected by season, plant development stage and growing environment, so that reference and basis are provided for researches of rapid identification of robinia pseudoacacia polyploids, new variety protection, genetic diversity analysis evaluation and genetic breeding. By adopting the primer taken as an example in the invention, the detection rate of polyploids in 343 robinia pseudoacacia clones is up to 85.42%, thus sufficiently indicating high efficiency and applicability of the SSR primer, and verifying that multiple robinia pseudoacacia polyploids generating natural variation really exist in the natural world.

Description

technical field [0001] The invention relates to the technical field of SSR molecular markers, in particular to primers for SSR molecular markers of Robinia pseudoacacia, and also relates to the application of the primers in identifying polyploids of Robinia pseudoacacia. Background technique [0002] Robinia pseudoacacia L. is a broad-leaved tree of the genus Robinia in the leguminous family (Leguminosae). It is resistant to drought, barrenness, mild salinity, pollution, and natural renewal. Pioneer tree species with high ornamental, nectar and feeding value. Robinia locust wood is tough, impact-resistant, decay-resistant, and resistant to water and humidity. It is an important material for construction, ore pillars, vehicles, and shipbuilding. Robinia locust wood has a high calorific value and a slow burning speed. It is a high-quality fuel wood and an excellent biomass energy tree species. Black locust is native to North America, and the worldwide introduction and domesti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 毛秀红荀守华王翠艳张元帅李双云李红静杨庆山孙百友闫少波
Owner SHANDONG FOREST SCI RES INST
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