Fragment combination, specific primer and application for identification of Rosaceae plant species

A species identification, Rosaceae technology, applied in the determination/inspection of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc.

Active Publication Date: 2021-08-03
INST OF BOTANY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, no effective identification method for Rosaceae plants has been reported.

Method used

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  • Fragment combination, specific primer and application for identification of Rosaceae plant species
  • Fragment combination, specific primer and application for identification of Rosaceae plant species
  • Fragment combination, specific primer and application for identification of Rosaceae plant species

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0126] Example 1, Screening Gene Loci and Primer Design for Identification of Rosaceae Plants

[0127] Collect a large number of measured Rosaceae chloroplast gene fragments and nuclear gene fragments to screen the initial data set of hypervariable gene fragments in Rosaceae plants, group the data sets according to gene fragments, and retain gene fragments containing at least 5 species, and use MAFFT software for comparison sequence, and then use BioEdit software to manually adjust the comparison results, and evaluate each gene fragment through the written python script. Finally, 7 gene fragments and 8 gene loci (matK, ITS-1, ITS-2, rbcL, trnH-psbA, atpB-rbcL, trnL, sbeI) were selected for the identification of Rosaceae plants.

[0128] Eight pairs of specific primers for the identification of Rosaceae plants were designed for the eight gene loci, and the specific information is shown in Table 1.

[0129] Table 1 Primer information

[0130]

Embodiment 2

[0131] Embodiment 2, establishment of detection method

[0132] 1. Extract the genomic DNA of the sample to be tested.

[0133] 2. Use the genomic DNA extracted in step 1 as a template, and use the primer pairs in Table 1 to perform PCR amplification to obtain PCR amplification products. The core codes matK and rbcL of the plant DNA standard barcode and the supplementary barcodes ITS and trnH- The primers corresponding to psbA are used for identification. If the primers corresponding to the above four barcodes cannot identify the species, other primers can be selected. For simple groups, a single or a combination of a small number of primer pairs can be selected for identification, and for complex groups, multiple primer pairs are required for joint identification. .

[0134] PCR amplification system (10μl): 10×PCR Buffer (Mg 2+ ) 1μl, 2mM dNTP 1μl, upstream primer 0.5μl, downstream primer 0.5μl, genomic DNA 1μl, 5U / μl Taq DNA polymerase 0.1μl, ddH 2 O 5.9 μl.

[0135] Eac...

Embodiment 3

[0141] Embodiment 3, actual detection

[0142] The DNA of all the samples in this example can be obtained from the Chinese Plant DNA Bank.

[0143] 1. Identify whether Rosa basilisk is a rare and endangered species

[0144] Samples to be tested: BOP214501 (Rose Rosaceae native to Lushun, used to identify whether it is a rare and endangered species).

[0145] The detection was carried out according to the method of Example 2, and the primer pair composed of Ros-ITS-1-F and Ros-ITS-1-R in Table 1 was used for PCR amplification. The sequencing result of the target fragment is shown in sequence 17 of the sequence listing.

[0146] The result is as figure 1 The topology is shown.

[0147] The results showed that Rosa basilicum escaped from European rose genetic resources.

[0148] 2. Cherry Blossom Identification

[0149] Sample to be tested: a dry leaf of an unknown cherry blossom, numbered BOP204816.

[0150] The detection was carried out according to the method of Example...

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Abstract

The invention discloses a fragment combination, a specific primer and an application thereof for identification of Rosaceae plant species. The primer combination provided by the present invention is composed of single-stranded DNA shown in sequence 1-sequence 16 of the sequence listing. Utilizing the specific primers provided by the invention, a general kit for identifying Rosaceae plant species can be developed to promote the application of plant DNA barcodes in social services.

Description

technical field [0001] The invention relates to a fragment combination, a specific primer and an application thereof for identification of Rosaceae plant species. Background technique [0002] Rosaceae is an economically important angiosperm, a large number of fruits (such as apples, pears, hawthorns, strawberries, peaches, plums, plums, apricots, cherries, etc.) and ornamental plants (such as plum blossoms, peach blossoms, cherry blossoms, roses, roses, Sorbus, etc.) belong to this family. In the subtropical and temperate regions of the northern hemisphere, Rosaceae is one of the large families. There are about 2800-3500 species in the world, and about 950 species in my country, more than half of which are endemic to my country. Issues such as counterfeiting and false crowns will bring about some economic disputes. For example, the cherries in our market are actually two different species: cherries (Prunus pseudocerasus) and European sweet cherries (C.avium). The more exp...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/11C12Q1/6895
CPCC12Q1/6895
Inventor 周世良杨雪莹陈荀张稳
Owner INST OF BOTANY CHINESE ACAD OF SCI
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