Specific primer pair based on rbcL gene and used for identifying land plant species and application thereof

A specific primer pair and species identification technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve problems such as loss and achieve good amplification effects

Inactive Publication Date: 2013-09-18
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, due to the loss of accD, ndhJ and ycf5 in some taxa, it was excluded in the later verification

Method used

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  • Specific primer pair based on rbcL gene and used for identifying land plant species and application thereof
  • Specific primer pair based on rbcL gene and used for identifying land plant species and application thereof
  • Specific primer pair based on rbcL gene and used for identifying land plant species and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Embodiment 1, the design of universal primer

[0017] The rbcL gene is located in the large single copy region (LSC) of the chloroplast genome (cpDNA), between the atpB and accD genes. The rbcL gene in higher plants does not contain introns, has a length of 1428bp, and encodes a total of 476 amino acids. Previous research work has shown that the rbcL gene has the characteristics of good versatility, easy amplification, and easy comparison. It is a very important gene in both molecular phylogenetic research and DNA barcode research. As one of the core DNA barcodes of plants, the rbcL gene can solve and improve the versatility of rbcL primers, and the design can be used in major plant groups such as angiosperms, gymnosperms, ferns, bryophytes, green algae and charophytes. Universal primers will promote the application of rbcL gene in phylogenetic studies and DNA barcoding.

[0018] The rbcL gene is the earliest molecular marker used in molecular systematics. In Genbank,...

Embodiment 2

[0024] Embodiment 2, the application of universal primer

[0025] There are 178 angiosperm families and 178 samples (one sample per family). There are 11 families and 11 samples of gymnosperms (one sample for each family). There are 33 families of ferns and 33 samples (one sample is selected for each family). There are 24 families and 24 samples of bryophytes (one sample is selected for each family).

[0026] A total of 178+11+33+24=246 samples (see Table 1).

[0027] 1. Extract the genomic DNA of the plant to be tested.

[0028] 2. Using the genomic DNA extracted in step 1 as a template, perform PCR amplification with a primer pair composed of rbcL-Forward Primer and rbcL-Reverse Primer to obtain a PCR amplification product.

[0029] PCR amplification system (10ul): 10×PCR Buffer (Mg 2+ ) 1ul, 2mM dNTP1ul, 5uM rbcL-Forward Primer0.5ul, 5uM rbcL-Reverse Primer0.5ul, 5ng-50ng / ul genomic DNA1ul, 5U / ul Taq DNA polymerase0.1ul, ddH 2 O5.9ul.

[0030] The instrument for PCR ...

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Abstract

The invention discloses a specific primer pair based on a rbcL gene and used for identifying land plant species and an application thereof. The specific primer pair provided by the invention is composed of a single stranded DNA A and a single stranded DNA B, wherein the single stranded DNA A is 15-40bp and comprises a DNA fragment of the DNA fragments as shown by a sequence 1 of a sequence table; and the single stranded DNA B is 15-40bp and comprises a DNA fragment of the DNA fragments as shown by a sequence 2 of the sequence table. The single stranded DNA A can be the DNA fragment as shown by the sequence 1 of the sequence table; and the single stranded DNA B can the DNA fragment as shown by the sequence 2 of the sequence table. The specific primer pair provided by the invention can be used for developing a universal kit, effectively identifying the land plant species and prompting the development and social service of plant DNA bar code.

Description

technical field [0001] The invention relates to a pair of specific primers based on rbcL gene for identification of terrestrial plant species and application thereof. Background technique [0002] DNA barcoding technology (DNA barcoding) refers to the use of a standard, sufficiently variable, easy-to-amplify, and relatively short DNA sequence fragment as a marker to achieve rapid, accurate, and automatic identification of species. The emergence of this technology has supplemented traditional taxonomic methods, and can solve species that cannot be identified by traditional taxonomic methods. In addition, it can also help to discover new species and hidden species, and can also be applied to ethnic medicine (such as Mongolian medicine) development and utilization. [0003] It was Paul Hebert, a professor at the University of Guelph in Canada and a member of the Royal Society of Canada, who first extended molecular methods to the entire biological taxonomy application and prop...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 周世良董文攀程涛李长昊徐超
Owner INST OF BOTANY CHINESE ACAD OF SCI
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