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A primer pair for amplifying the barcode of Liriodendron tulipifera and the identification method of Liriodendron tulipifera and its provenance

A technology of barcode primers and amplification primers, which is applied in biochemical equipment and methods, recombinant DNA technology, DNA/RNA fragments, etc., can solve the problems of insufficient identification dimension and large variation of morphological markers in Liriodendron tulipifera, and achieve High accuracy and simple operation

Active Publication Date: 2022-04-12
INST OF BIOLOGICAL RESOURCES JIANGXI ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Traditional classification and identification mostly rely on differences in phenotypic characteristics, but morphological markers vary greatly, and usually require complete plant tissue parts; although the emergence of SSR molecular markers in recent years has solved the problem of Chinese tulip tree, North American tulip tree, and North American to a certain extent. Part of the identification of Liriodendron tulipifera and hybrid Liriodendron tulipifera, but the identification dimension within the species of Liriodendron tulipifera is not enough, and multiple pairs of primers are needed in combination

Method used

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  • A primer pair for amplifying the barcode of Liriodendron tulipifera and the identification method of Liriodendron tulipifera and its provenance
  • A primer pair for amplifying the barcode of Liriodendron tulipifera and the identification method of Liriodendron tulipifera and its provenance
  • A primer pair for amplifying the barcode of Liriodendron tulipifera and the identification method of Liriodendron tulipifera and its provenance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] The entire distribution area of ​​the goose is to collect North American Goose, and the goose, a total of 120 samples (sample number 1-120). Related information such as sample sources is shown in Table 1.

[0044] These samples were identified using the DNA strip primer pair and method of the present invention.

[0045] details as follows:

[0046] 1) Extract the sample tender vesicular genomic DNA with improved CTAB method.

[0047] 2) The amplification product was obtained using the DNA strip primer to obtain PCR amplification with the total genome DNA obtained. The PCR amplification system is 20 μl, including 2 × Taq PCR MASTER MIX 10 μL, 20 ~ 40 ng / l all genomic DNA 2.0 μL, 10 μmol / l amplification primer 0.6 μL, 10 μmol / L, downstream amplification primer 0.6 μL, DDH 2 O 6.8 μL; PCR amplification procedures include: 95 ° C for 4 min; 95 ° C denaturation 40S, 59 ° C annealing 40S, 72 ° C extension 40s, 33 cycles; 72 ° C for 6 min.

[0048] 3) Single as a sequencing ...

Embodiment 2

[0052] Collect the 43 sources of the pendant of the goose 's all parties (sample numbers 13 to 164), cover the maximum distribution area of ​​the goose, and the sample source, etc. All individuals collect young leaves and stored with silica gel.

[0053] 1) Total DNA extraction of Goose in the Goose using the improved CTAB method.

[0054] 2) The amplification product was obtained using the DNA strip primer to obtain PCR amplification with the total genome DNA obtained. The PCR amplification system is 20 μl, including 2 × Taq PCR MASTER MIX10 μL, 20 ~ 40 ng / l all genomic DNA 2.0 μL, 10 μmol / L amplification primer 0.6 μL, 10 μmol / L of downstream amplification primers 0.6 μL, DDH 2 O 6.8 μL; PCR amplification procedures include: 95 ° C for 4 min; 95 ° C denaturation 40S, 59 ° C annealing 40S, 72 ° C extension 40s, 33 cycles; 72 ° C for 6 min.

[0055] 3) Single as a sequencing primer in SEQ ID NO.1, commissioning biotech companies in one-way sequencing, obtain sequencing resul...

Embodiment 3

[0059] Sample source and operation steps are in Example 2.

[0060] By sequencing and sequence alignment, it was found that from 375 bp, 8 A base repeats were started from 375 bp, and the tropical source standard sequence SEQ ID NO.6 of the foop of the present invention was identified as a tropical. Sources, the remaining samples were all three A base repetitions from 375 bp. No. 24 ~ 26 and numbered samples with DNA barcode sequences (SEQ ID NO.5 and SEQ ID NO.6) are met in 320 ~ 395 bp regional sequence. Figure 4 The source of controls found that all samples from 13 to 26 come from the tropics, and the remaining samples are from the subtropical region. Therefore, the identification accuracy of the barcode of the present invention is 100%.

[0061] As can be seen from the above embodiment, the method of identifying the geese and its geese from the identification method of the goose crowd and the identification method of the goose. The present invention provides strong support for...

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Abstract

The invention provides a primer pair for amplifying the DNA barcode of Liriodendron and an identification method for Liriodendron and its provenance, belonging to the technical field of plant molecular identification. The primer pair for amplifying the DNA barcode of Liriodendron includes upstream amplification An amplification primer and a downstream amplification primer; the sequence of the upstream amplification primer is shown in SEQ ID No.1; the sequence of the downstream amplification primer is shown in SEQ ID No.2. Liriodendron tulipifera identification method provided by the invention comprises the following steps: 1) extracting the whole genome DNA of the sample to be detected; 2) using the whole genome DNA as a template, performing PCR amplification with the DNA barcode primer pair to obtain an amplified product 3) Sequencing the amplified product, obtaining the specific sequence information of the amplified product, comparing the specific sequence information with the standard sequence of the Liriodendron tulipifera DNA barcode, and determining the tree species and regional provenance of the sample to be tested, the The method is simple in operation and high in accuracy.

Description

Technical field [0001] The present invention belongs to the field of plant molecules identification, and in particular, the present invention relates to an identification method of primer pairs and geese bullion and its geese. Background technique [0002] Liriodendron Chinense (Hemsl.) Sarg.) Magnoliaceae's geese is a geese, for many years of deciduous tree species. Goose palm leaves, ornamental value is high; fast-growing, wood is straight, material fine, and is a good material for tree species and potential energy plants. The geese is widely distributed in the Temperary Treatment area of ​​the Northern Hemisphere, followed by the climate turmoil in the third discipline, currently distributing in small parts of East Asia and North America, and differentiates into geese, northern geese ( L.Tulipifera). North American Goose is distributed in the broad-leaved forest area in eastern United States, which is the advantages and pioneer species of the southern forest area of ​​Abarachi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/156
Inventor 杨爱红刘淑娟刘立盘钟永达刘腾云周华吴照祥李彦强余发新
Owner INST OF BIOLOGICAL RESOURCES JIANGXI ACAD OF SCI
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