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Molecular distinctness labeling primers for aster dendrobium and detection method

A technology for labeling primers and specificity, which is used in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc. It can solve the problems of long cycle, poor repeatability and low specificity, and achieve the effect of simple method.

Inactive Publication Date: 2016-11-09
ZHEJIANG FORESTRY ACAD
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AI Technical Summary

Problems solved by technology

[0004] The identification of Dendrobium species is mainly based on appearance characteristics, but due to the long period of identification of many morphological characters, the impact of the environment, and the increasing number of varieties, the identification of varieties is becoming more and more difficult.
Since this century, some PCR-based molecular marker technologies such as RAPD (Random Amplified Polymorphic DNA, Random Amplified Polymorphic DNA), ISSR (Inter-Simple Sequence Repeat, Simple Sequence Repeat Interval Amplified Polymorphism) and SRAP ( Sequence-related amplified polymorphism) has been successively used to study the genetic diversity and genetic distance of Dendrobium germplasm resources, but there are few studies on the linkage between molecular markers and Dendrobium traits, and molecular identification between Dendrobium varieties. Moreover, these molecular markers are either universal random primers or a random combination of designed primers, the PCR amplification pattern is not only complicated, the repeatability is poor, and the specificity is not high, so it is not suitable for variety identification
SCAR (Sequence Characterized Amplified Region) marker was proposed by Paran and Michelmore on the basis of RAPD in 1993. It is based on the sequencing of specific RAPD fragments, and a pair of 18-24 bases is designed according to the sequences at both ends. Primers are achieved by specific amplification at a higher annealing temperature, and the use of specific primers eliminates the competition between random primer binding sites, so it is a very stable molecular marker and has a wide range of applications. Rapid, simple, and low-cost characteristics, so far, there is no research report on the application of SCAR markers in the identification of Dendrobium species

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  • Molecular distinctness labeling primers for aster dendrobium and detection method
  • Molecular distinctness labeling primers for aster dendrobium and detection method

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Embodiment 1

[0037] (1) Extraction of genomic DNA of Dendrobium species: Take 0.01g of young leaves of Dendrobium species to be tested, add liquid nitrogen to grind thoroughly, and use SDS-CTAB method to extract genomic DNA. After repeated extractions, extract the genome of Dendrobium Crude DNA extract. The crude DNA was purified by Magabio Nucleic Acid Purification Kit (Bioer, Hangzhou, China), and detected by 1.5% agarose gel electrophoresis and DNA / RNA UV spectrophotometer (GeneQuant Pro, GE Healthcare). Integrity, Purity and Concentration. OD 260 / OD 280 DNA samples >1.8 were used for subsequent PCR amplification. DNA extracts were stored in a -20°C refrigerator for later use.

[0038] (2) Design specific PCR amplification primers. The sequence of the primer pair is: 5'-AGCGGCCGCGCCTTCC-3' and the downstream primer 5'-GGAAGGCGCGGCCGCT-3', synthesized by Hangzhou Qingke Engineering Technology Co., Ltd.

[0039] (3) PCR amplification of SCAR molecular markers:

[0040] The composit...

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Abstract

The invention relates to molecular distinctness labeling primers for aster dendrobium and a method capable of rapidly identifying aster dendrobium. According to the primers, the sequence of the upstream primer is 5'-AGCGGCCGCGCCTTCC-3', and the sequence of the downstream primer is 5'-GGAAGGCGCGGCCGCT-3'. The molecular distinctness labeling primers for aster dendrobium and the method capable of rapidly identifying aster dendrobium have the advantages that the molecular distinctness labeling primers can carry out rapid early-stage identification on aster dendrobium, and the method is simple, fast and accurate, and is a molecular means which cannot be replaced by the method of distinguishing dendrobium varieties with the apparent characteristics.

Description

(1) Technical field [0001] The invention relates to a molecular specific marker primer of Dendrobium asteroides and a method for identifying and detecting Dendrobium asteraceae using the primers. (2) Background technology [0002] Dendrobium (D.orchids), also known as Dendrobium orchid, is a plant of the genus Dendrobium (Orchidaceae), widely distributed in China, Japan, the Philippines, Thailand, India, Malaysia, Australia and New Zealand in Asia and Oceania. There are more than 1,000 species in my country, and there are 74 species and 2 varieties in China. As an excellent medicinal plant, Dendrobium has high medicinal and health value. It was first seen in "Shan Hai Jing", and its medicinal use was first recorded in "Shen Nong's Materia Medica", and was later recorded in "Compendium of Materia Medica" and other successive dynasties. , has the effects of "mainly harming, eliminating numbness, lowering qi, tonifying the five Zangs, strengthening yin, thickening the stomach, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
CPCC12Q1/6895
Inventor 童晓青王丽玲秦川王衍彬刘本同黄旭波方茹钱华
Owner ZHEJIANG FORESTRY ACAD
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