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Marker, primer, kit and identifying method for cordate houttuynia molecular identification

A technology for molecular identification and Houttuynia cordata, applied in the field of plant molecular biology, can solve problems such as no molecular identification of Houttuynia cordata, no Houttuynia cordata report, etc., and achieves the reduction of manual sequencing labor, easy standardized analysis, and easy plant identification. Effect

Inactive Publication Date: 2015-05-06
HUNAN AGRICULTURAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] But so far, there is no application of DNA barcoding technology for molecular identification of Houttuynia cordata, and there is no report of a special barcode for Houttuynia cordata

Method used

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  • Marker, primer, kit and identifying method for cordate houttuynia molecular identification
  • Marker, primer, kit and identifying method for cordate houttuynia molecular identification
  • Marker, primer, kit and identifying method for cordate houttuynia molecular identification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Embodiment 1: the process that the inventive method implements

[0027] (1) Preparation of sample materials: take about 10 g of young leaves of Houttuynia cordata and other plants of the same family.

[0028] (2) Genomic DNA extraction: use CTAB method to extract genomic DNA, grind freeze-dried tobacco leaves into powder, add 65°C preheated CTAB extract, keep warm at 65°C for more than 45min, and shake gently occasionally → 12000r / After centrifuging at room temperature for 20 minutes, take the supernatant → add an equal volume of chloroform:isoamyl alcohol (24:1), mix gently for more than 15 minutes, centrifuge at 12000r / min for 20 minutes at room temperature, take the supernatant and transfer it to a new centrifuge tube → add Pre-cooled isopropanol, shake gently for 5min, centrifuge at 8000r / min for 10min at room temperature and remove the supernatant → extract the precipitate with 75% ethanol and 10mmol / L KAC for 2-3 times (centrifuge at room temperature for 10min at...

Embodiment 2

[0034] Example 2: Difference analysis of material DNA barcodes after the method of the present invention is implemented

[0035]Apply the primers of the present invention to carry out PCR amplification on the material, and obtain fragments with sequence lengths of about 470bp, which have little variation (between 0.0% and 0.16%) among Houttuynia cordata species from different origins, and The information site in Houttuynia species is very stable; and the degree of variation between species is large (between 6.2%-9.1%), so this sequence is suitable as a barcode sequence for identifying Houttuynia plants, and it is easy Distinguish Houttuynia cordata from different species of plants such as white bracts.

[0036]

[0037]

[0038] The above is the alignment of the amplified base sequences of Houttuynia cordata and Pyllium baculum. The shades represent different parts of the two sequences, and the underlines represent the primer positions.

[0039]

[0040]

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Abstract

The invention relates to a marker, a primer, a kit and an identifying method for cordate houttuynia molecular identification, and belongs to the field of plant molecular biology. The method comprises the following steps: comparing a DNA fragment amplified by PCR of a to-be-identified sample with a DNA bar code of cordate houttuynia plant by sequencing; identifying that the to-be-identified sample is cordate houttuynia when the similarity between an amplification product and a base sequence as shown in the cordate houttuynia DNA bar code is over 98%. The identifying method overcomes a technical problem that a sensory evaluation method is difficult to identify whether the cordate houttuynia is real or not, and especially can be used for quickly and effectively distinguishing the cordate houttuynia from counterfeit-species gymnotheca involucrate.

Description

technical field [0001] The invention belongs to the field of plant molecular biology, in particular to a marker, primer, kit and identification method for molecular identification of houttuynia cordata. Background technique [0002] Houttuynia cordata Thunb (Houttuynia cordata Thunb) belongs to the genus Houttuynia cordata and is a perennial herbaceous plant. Commonly known as pear root, pig's nostril, stinky grass, Qincao, etc., because of the fishy smell after the stems and leaves are crushed, it is named Houttuynia cordata. Houttuynia cordata not only can be eaten as vegetables with tender stems, leaves and underground stems, but also the whole plant can be used fresh or dried as medicine. It has been officially identified by the Ministry of Health as a resource with great development potential that is "both medicine and food". One of them is gaining more and more attention. [0003] Due to the sharp increase in the demand for Houttuynia cordata at home and abroad, the ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6869C12Q1/6895
Inventor 钟军熊兴耀
Owner HUNAN AGRICULTURAL UNIV